L002: Cellular components and organizationPeter TakizawaDepartment of Cell Biology

•Course organization

•Why cell biology

•How we study and think about cells

•Plasma membrane

Molecules to Systems comprises Biochemistry, Cell Biology and Physiology.

Biochemistry Cell Biology Physiology

Cell biology is part of the molecules to systems integrated curriculum, along with biochemistry and physiology. Each is its own, separate course with different course directors and individual curriculum. Because the content of the courses had much in common, we decided several years ago to try to integrate the content between the courses. That means, the course directors of each course, Susan Baserga of Biochemistry and Emil Boulpaep of Physiology, meet to discuss the content of our courses to ensure that we adequately cover all the relevant topics while avoiding excessive redundancy between the courses. In addition, the timing of the topics between the courses has been coordinated so that we present related content during the same time frame.

Molecules to Systems comprises Biochemistry, Cell Biology and Physiology.

Biochemistry Cell Biology Physiology

Cell biology is part of the molecules to systems integrated curriculum, along with biochemistry and physiology. Each is its own, separate course with different course directors and individual curriculum. Because the content of the courses had much in common, we decided several years ago to try to integrate the content between the courses. That means, the course directors of each course, Susan Baserga of Biochemistry and Emil Boulpaep of Physiology, meet to discuss the content of our courses to ensure that we adequately cover all the relevant topics while avoiding excessive redundancy between the courses. In addition, the timing of the topics between the courses has been coordinated so that we present related content during the same time frame.

Lectures in Biochemistry, Cell Biology and Physiology are ordered with each other.

Date Start Time End Time Lecture Title Format Room FacultyAug 29, 2012 11:00 AM 11:50 AM BC1_LE001 Intro: Every Atom Counts Lecture Hope 110 Baserga, SAug 29, 2012 12:00 PM 12:50 PM CB1_LE002 Cellular Components and Organization Lecture Hope 110 Takizawa, PAug 30, 2012 9:00 AM 9:50 AM BC1_LE003 Proteins Lecture Hope 110 Engelman, DAug 30, 2012 10:00 AM 10:50 AM CB1_LE004 Protein Localization Lecture Hope 110 Takizawa, PAug 30, 2012 11:00 AM 12:20 PM PY1_WS000 Organizational meeting Workshop TBA Boulpaep, EAug 31, 2012 9:00 AM 9:50 AM BC1_LE005 Lipids Lecture Hope 110 Engelman, DAug 31, 2012 10:00 AM 10:50 AM PY1_LE006 Fluid compartments/resting potentials Lecture Hope 110 Boulpaep, E

Sep 4, 2012 9:00 AM 9:50 AM PY1_LE007 Membrane transport & regulation Lecture Hope 110 Boulpaep, E

Because of the integration created between the courses, the activities of the courses were arranged into one master schedule that continues today. Consequently, the lectures in each course are numbered sequentially with respect to each other. That means the lecture number for a particular course’s lecture refers to the number of the lecture in Molecules to Systems curriculum and not in the course. For example, this is the first lecture in Cell Biology but is numbered 002 because it follows a biochemistry lecture. My second lecture is numbered 004. This made more sense when we were a paper-based curriculum and students were given a spreadsheet listing all the activities in MSIC.

Lectures in Biochemistry, Cell Biology and Physiology are ordered with each other.

Date Start Time End Time Lecture Title Format Room FacultyAug 29, 2012 11:00 AM 11:50 AM BC1_LE001 Intro: Every Atom Counts Lecture Hope 110 Baserga, S

Aug 29, 2012 12:00 PM 12:50 PM CB1_LE002 Cellular Components and Organization Lecture Hope 110 Takizawa, PAug 30, 2012 9:00 AM 9:50 AM BC1_LE003 Proteins Lecture Hope 110 Engelman, DAug 30, 2012 10:00 AM 10:50 AM CB1_LE004 Protein Localization Lecture Hope 110 Takizawa, PAug 30, 2012 11:00 AM 12:20 PM PY1_WS000 Organizational meeting Workshop TBA Boulpaep, EAug 31, 2012 9:00 AM 9:50 AM BC1_LE005 Lipids Lecture Hope 110 Engelman, DAug 31, 2012 10:00 AM 10:50 AM PY1_LE006 Fluid compartments/resting potentials Lecture Hope 110 Boulpaep, E

Sep 4, 2012 9:00 AM 9:50 AM PY1_LE007 Membrane transport & regulation Lecture Hope 110 Boulpaep, E

Because of the integration created between the courses, the activities of the courses were arranged into one master schedule that continues today. Consequently, the lectures in each course are numbered sequentially with respect to each other. That means the lecture number for a particular course’s lecture refers to the number of the lecture in Molecules to Systems curriculum and not in the course. For example, this is the first lecture in Cell Biology but is numbered 002 because it follows a biochemistry lecture. My second lecture is numbered 004. This made more sense when we were a paper-based curriculum and students were given a spreadsheet listing all the activities in MSIC.

Cell Biology

Lectures

Histology

ClinicalCorrelations

Bench to Bedside

Mol. Basis Disease

Electives

Cell Biology comprises a variety of activities that discuss basic science and disease.

The Cell Biology course proper consists of three distinct activities: lectures, histology labs and clinical correlations. In addition, there are two electives that are associated with Cell Biology: molecular and cellular basis of disease and bench to bedside.

Lectures will discuss the principles and concepts of modern cellular and molecular biology, focusing on the systems and mechanisms that allow cells to survive and perform specific functions in our bodies. The first part of the course will discuss the systems and mechanisms that are common to most cells. The second part will discuss how the different types of cells in our bodies, utilize and modify those systems to perform specific biological functions.

Histology examines the structure and functions of cells and how cells form tissues and organs. Histology places the cellular mechanisms presented in lecture into the context of cell and tissue structure. Histology also demonstrates how the organization of cell and tissues allows organs to perform the physiological functions.

Clinical correlations introduce students to clinical topics and medical terminology and demonstrate connections between basic science and disease. These presentations by physician-scientists, who are leaders in their fields, will sometimes include patients. You will notified when a patient is present.

Molecular and cellular basis of disease emphasizes connections between diseases and basic science using lecture and seminar format. It is designed for students who are committed to or considering careers in medical research. The elective will explore scientific topics and their impact on human disease in detail. First meeting 9/10 at 4 PM SHM-428.

Bench to Bedside comprises a series of seminars that show students how questions derived from patients can help them learn more about biological and pathological processes. The seminars focus on pediatrics and diseases and medical problems that affect infants and children. Students select a faculty preceptor and topic and then observe a patient with the preceptor. Students will then present the patient to the group and discuss questions that arise in learning about the diseases affecting the patient.

Both electives are required for MD-PhD students and any student considering joining the MD-PhD program.

Cells are the fundamental units of life.

Cells are the fundamental units of life. What that means is that cells are the smallest unit capable of growth, replication and ability to adapt to environment. These principles are exhibited by the many different types of single cells organisms, such as bacteria and yeast, that can grow and divide on their own and respond to changes in the environment to survive.

But even cells from multicellular organisms, like ourselves, can survive as individual cells and under appropriate conditions grow and divide. This second image shows human cells growing as individual cells in culture. In 1951, Henrietta Lacks was diagnosed with cervical cancer and a portion of the cancer was removed and the cells cultured in petri dishes. Descendants from these cells are growing today and are widely used in biomedical research.

The fact that cells can grow and adapt to environmental changes makes them fascinating subjects of study for those interested in biology and science. But why study cell biology in medical school?

Cells are the fundamental units of life.

Cells are the fundamental units of life. What that means is that cells are the smallest unit capable of growth, replication and ability to adapt to environment. These principles are exhibited by the many different types of single cells organisms, such as bacteria and yeast, that can grow and divide on their own and respond to changes in the environment to survive.

But even cells from multicellular organisms, like ourselves, can survive as individual cells and under appropriate conditions grow and divide. This second image shows human cells growing as individual cells in culture. In 1951, Henrietta Lacks was diagnosed with cervical cancer and a portion of the cancer was removed and the cells cultured in petri dishes. Descendants from these cells are growing today and are widely used in biomedical research.

The fact that cells can grow and adapt to environmental changes makes them fascinating subjects of study for those interested in biology and science. But why study cell biology in medical school?

Cells are the fundamental units of life.

Cells are the fundamental units of life. What that means is that cells are the smallest unit capable of growth, replication and ability to adapt to environment. These principles are exhibited by the many different types of single cells organisms, such as bacteria and yeast, that can grow and divide on their own and respond to changes in the environment to survive.

But even cells from multicellular organisms, like ourselves, can survive as individual cells and under appropriate conditions grow and divide. This second image shows human cells growing as individual cells in culture. In 1951, Henrietta Lacks was diagnosed with cervical cancer and a portion of the cancer was removed and the cells cultured in petri dishes. Descendants from these cells are growing today and are widely used in biomedical research.

The fact that cells can grow and adapt to environmental changes makes them fascinating subjects of study for those interested in biology and science. But why study cell biology in medical school?

Cells are the fundamental units of life.

Cells are the fundamental units of life. What that means is that cells are the smallest unit capable of growth, replication and ability to adapt to environment. These principles are exhibited by the many different types of single cells organisms, such as bacteria and yeast, that can grow and divide on their own and respond to changes in the environment to survive.

But even cells from multicellular organisms, like ourselves, can survive as individual cells and under appropriate conditions grow and divide. This second image shows human cells growing as individual cells in culture. In 1951, Henrietta Lacks was diagnosed with cervical cancer and a portion of the cancer was removed and the cells cultured in petri dishes. Descendants from these cells are growing today and are widely used in biomedical research.

The fact that cells can grow and adapt to environmental changes makes them fascinating subjects of study for those interested in biology and science. But why study cell biology in medical school?

We are made entirely of cells and material produced by cells.

We are made entirely of cells and the material that cells produce. We could dissect out someone’s liver, an organ that helps us digest our food, metabolize nutrients, breakdown toxins and is essential to our survival. We can take a section from that liver and look at it under a microscope. We can zoom in on the section to reveal...cells. In fact, it’s almost all cells. So all the critical physiological functions that are performed by the liver, that are essential for our survival, are performed by these cells. When these cells stop working, which can happen for a variety of genetic and environmental reasons, the liver works less efficiently and could eventually compromise our ability to survive. So to understand how we grow and survive, we have to understand to some degree how cells work. Further, to understand and treat disease, we need to understand how the failures of cells negatively affects our physiology and consequently our ability to survive.

For example, the inability of these cells to process triglycerides leads to an accumulation of fat within the cytoplasm of the cells, called steatosis. This reduces the function of the cells and if allowed to progress will lead to inflammation and cell death and liver failure.

We are made entirely of cells and material produced by cells.

We are made entirely of cells and the material that cells produce. We could dissect out someone’s liver, an organ that helps us digest our food, metabolize nutrients, breakdown toxins and is essential to our survival. We can take a section from that liver and look at it under a microscope. We can zoom in on the section to reveal...cells. In fact, it’s almost all cells. So all the critical physiological functions that are performed by the liver, that are essential for our survival, are performed by these cells. When these cells stop working, which can happen for a variety of genetic and environmental reasons, the liver works less efficiently and could eventually compromise our ability to survive. So to understand how we grow and survive, we have to understand to some degree how cells work. Further, to understand and treat disease, we need to understand how the failures of cells negatively affects our physiology and consequently our ability to survive.

For example, the inability of these cells to process triglycerides leads to an accumulation of fat within the cytoplasm of the cells, called steatosis. This reduces the function of the cells and if allowed to progress will lead to inflammation and cell death and liver failure.

We are made entirely of cells and material produced by cells.

We are made entirely of cells and the material that cells produce. We could dissect out someone’s liver, an organ that helps us digest our food, metabolize nutrients, breakdown toxins and is essential to our survival. We can take a section from that liver and look at it under a microscope. We can zoom in on the section to reveal...cells. In fact, it’s almost all cells. So all the critical physiological functions that are performed by the liver, that are essential for our survival, are performed by these cells. When these cells stop working, which can happen for a variety of genetic and environmental reasons, the liver works less efficiently and could eventually compromise our ability to survive. So to understand how we grow and survive, we have to understand to some degree how cells work. Further, to understand and treat disease, we need to understand how the failures of cells negatively affects our physiology and consequently our ability to survive.

For example, the inability of these cells to process triglycerides leads to an accumulation of fat within the cytoplasm of the cells, called steatosis. This reduces the function of the cells and if allowed to progress will lead to inflammation and cell death and liver failure.

We are made entirely of cells and material produced by cells.

We are made entirely of cells and the material that cells produce. We could dissect out someone’s liver, an organ that helps us digest our food, metabolize nutrients, breakdown toxins and is essential to our survival. We can take a section from that liver and look at it under a microscope. We can zoom in on the section to reveal...cells. In fact, it’s almost all cells. So all the critical physiological functions that are performed by the liver, that are essential for our survival, are performed by these cells. When these cells stop working, which can happen for a variety of genetic and environmental reasons, the liver works less efficiently and could eventually compromise our ability to survive. So to understand how we grow and survive, we have to understand to some degree how cells work. Further, to understand and treat disease, we need to understand how the failures of cells negatively affects our physiology and consequently our ability to survive.

For example, the inability of these cells to process triglycerides leads to an accumulation of fat within the cytoplasm of the cells, called steatosis. This reduces the function of the cells and if allowed to progress will lead to inflammation and cell death and liver failure.

How do we study cells?

• Microscopy

• Biochemistry

• Molecular genetics

There are a variety of methods and disciplines that examine cells. The most common approaches are microscopy, biochemistry and molecular genetics.

Microscopy is the foundation of cell biology.

Of the three, microscopy is the oldest. Ever since Leeuwenhoek described single-cell organisms in his hand-made microscope. Scientist have used microscopes to study the structure and organization of cells. With microscopes we can not only view individual cells, but we can look at the subcellular components of cells. We can watch the movement of those components over time. And more recently, we can watch the behavior of individual cells within living organisms.

Microscopy is the foundation of cell biology.

Of the three, microscopy is the oldest. Ever since Leeuwenhoek described single-cell organisms in his hand-made microscope. Scientist have used microscopes to study the structure and organization of cells. With microscopes we can not only view individual cells, but we can look at the subcellular components of cells. We can watch the movement of those components over time. And more recently, we can watch the behavior of individual cells within living organisms.

Microscopy is the foundation of cell biology.

Of the three, microscopy is the oldest. Ever since Leeuwenhoek described single-cell organisms in his hand-made microscope. Scientist have used microscopes to study the structure and organization of cells. With microscopes we can not only view individual cells, but we can look at the subcellular components of cells. We can watch the movement of those components over time. And more recently, we can watch the behavior of individual cells within living organisms.

Microscopy is the foundation of cell biology.

Of the three, microscopy is the oldest. Ever since Leeuwenhoek described single-cell organisms in his hand-made microscope. Scientist have used microscopes to study the structure and organization of cells. With microscopes we can not only view individual cells, but we can look at the subcellular components of cells. We can watch the movement of those components over time. And more recently, we can watch the behavior of individual cells within living organisms.

Biochemistry identifies the molecular pathways that are essential for cell viability and function.

The second major approach to studying cells is biochemistry. Ever since scientists discovered that they could crack open cells and recapitulate many of the process that occur in cells in test tubes, scientists have used this approach to define the biochemical pathways that occur in cells, such as glycolysis, and to identify the components that mediate these pathways.

Molecular genetics shows how changes in protein sequence affects cell function and viability.

Normal Cell Mutant Cell

More recently with the advent of molecular genetics and its ability to manipulate and sequence genes has allowed us to study how changes in the amino acids of individual proteins affect their functions. We can make mutations in key regions of a protein and then test how those mutations affect a given cellular activity.

Modern cell biology utilizes all three approaches (and more) to study cells.

Microscopy

Biochemistry MolecularGenetics

Today, cell biologist employ all three methods to study cells with the hope that we can understand how individual mutations lead to changes in protein and cell function

Mutations in muscle myosin affect cardiac muscle, leading to diseases of the heart.

Cel

l56

0

Figu

re4.

A3D

Rep

rese

ntat

ion

ofth

eA

ctin

–M

yosi

nC

ompl

ex,

with

Myo

sin

Por

tray

edin

the

Pos

t-P

ower

Str

oke

(Rig

orC

onfo

rmat

ion)

afte

rS

epar

atio

nfr

omA

ctin

One

myo

sin

head

and

two

actin

mon

omer

sar

edi

spla

yed

with

actin

–myo

sin

bind

ing

do-

mai

ns(g

reen

),A

TPbi

ndin

gdo

mai

nin

myo

sin

(yel

low

),an

dth

em

yosi

nes

sent

ial

(ora

nge)

and

regu

lato

ry(v

iole

t)lig

htch

ains

high

-lig

hted

.Th

ese

para

tion

ofac

tinan

dm

yosi

nha

sbe

enin

crea

sed

for

clar

ity.

Res

idue

sin

actin

and

myo

sin

that

are

mut

ated

inhy

per-

trop

hic

card

iom

yopa

thy

are

show

nin

blue

;re

sidu

esth

atar

em

utat

edin

dila

ted

card

io-

myo

path

yar

esh

own

inre

d(a

dapt

edfr

omR

aym

ent

etal

.,19

93;S

chro

eder

etal

.,19

93;

and

Sei

dman

and

Sei

dman

,200

0).

spec

ific

proc

esse

sth

atar

epe

rtur

bed

byth

ese

defe

cts.

1998

),an

dtr

opon

inT

(Obe

rst

etal

.,19

98;T

ardi

ffet

al.,

1998

)ha

vebe

enge

netic

ally

engi

neer

edin

toge

nom

esA

nim

alm

odel

sha

vebe

enpa

rtic

ular

lyva

luab

lebo

thbe

-ca

use

stab

lece

lllin

esof

myo

cyte

linea

ges

dono

texi

st,

ofm

ice

and

rabb

its(M

aria

net

al.,1

999)

.Th

ese

mod

els

exhi

bit

man

ych

arac

teris

tics

ofth

ehu

man

dise

ase,

in-

and

beca

use

trad

ition

alst

udie

sof

nong

enet

icfo

rms

ofca

rdia

chy

pert

roph

ypr

oduc

edby

surg

ical

orph

arm

aco-

clud

ing

card

iac

hype

rtro

phy

and

hist

opat

holo

gic

fea-

ture

sof

myo

cyte

disa

rray

with

incr

ease

din

ters

titia

lfi-

logi

cal

inte

rven

tions

indi

cate

the

impo

rtan

ceof

inte

-gr

ativ

eor

gan

phys

iolo

gyin

card

iac

rem

odel

ing.

Hum

anbr

osis

and

alte

red

card

iac

phys

iolo

gy.

Aco

nsis

tent

findi

ngfr

oman

alys

esof

thes

em

odel

sin

dica

tes

that

mut

atio

nsin

myo

sin

heav

ych

ain

(Gei

ster

fer-

Low

ranc

eet

al.,

1996

),m

yosi

nbi

ndin

gpr

otei

nC

(Yan

get

al.,

card

iac

phen

otyp

eis

not

the

cons

eque

nce

ofha

ploi

n-

Tabl

e1.

Gen

etic

and

Clin

ical

Het

erog

enei

tyof

Hum

anC

ardi

omyo

path

y

Locu

sA

dditi

onal

Phe

noty

peD

isea

seG

ene

Mut

atio

nTy

pe

Hyp

ertr

ophi

c

1q32

Non

eC

ardi

acTr

opon

inT

Mis

sens

e,D

elet

ions

,Spl

ice

Def

ects

2q31

Non

eTi

tinM

isse

nse

11p1

1N

one

Car

diac

Myo

sin

Bin

ding

Pro

tein

-CM

isse

nse,

Del

etio

ns,S

plic

eD

efec

ts14

q12

Non

eC

ardi

ac�

Myo

sin

Hea

vyC

hain

Mis

sens

e15

q14

Non

eC

ardi

acA

ctin

Mis

sens

e15

q22

Non

e�

Trop

omyo

sin

Mis

sens

e19

q13

Non

eC

ardi

acTr

opon

inI

Mis

sens

e3p

21S

kele

talM

yopa

thy

Ess

entia

lMyo

sin

Ligh

tC

hain

Mis

sens

e12

q23-

p21

Ske

leta

lMyo

path

yR

egul

ator

yM

yosi

nLi

ght

Cha

inM

isse

nse

7q3

Pre

-exc

itatio

n?

Dila

ted

1q32

Non

eC

ardi

acTr

opon

inT

Del

etio

ns1q

32N

one

?2q

11-q

22N

one

?2q

31N

one

?9q

13-q

22N

one

?14

q11

Non

eC

ardi

ac�

Myo

sin

Hea

vyC

hain

Mis

sens

e15

q14

Non

eC

ardi

acA

ctin

Mis

sens

e10

q21-

q23

Mitr

alVa

lve

Pro

laps

e?

1p1-

q21

Con

duct

ion

Dis

ease

Lam

inA

/CM

isse

nse

2q11

-q22

Con

duct

ion

Dis

ease

?3q

22-p

25C

ondu

ctio

nD

isea

se?

19q1

3.2

Con

duct

ion

Dis

ease

?2q

35S

kele

talM

yopa

thy

Des

min

Mis

sens

e6q

23S

kele

talM

yopa

thy

?X

p21

Ske

leta

lMyo

path

yD

ystr

ophi

nD

elet

ions

Xq2

8S

hort

Sta

ture

&N

eutr

open

iaTa

fazz

inD

elet

ions

,Spl

icin

gD

efec

ts6q

23-2

4S

enso

rineu

ralH

earin

gLo

ss?

MyosinShown in this example is myosin of that powers the contraction of muscle cells. The colored dots show individual amino acids which when changed alter the activity of myosin. We can correlate the changes in myosin sequence and activity with changes in the structure and function of cells and tissues. In this case, cardiac muscle cells that mediate contraction of the heart. The changes in cell function can be seen to alter the overall structure of organs, not the increased size of the muscle tissue in the heart with the mutation in muscle myosin and the decreased volume of the left ventricle. Finally, the changes in the structure of the heart compromise its activity leading to disease and often death. So how do we think about cells?

Mutations in muscle myosin affect cardiac muscle, leading to diseases of the heart.

Cell558

myocytes that characterize the normal myocardium (Fig-ure 2A) becomes distorted by hypertrophic growth ofthe myocytes, which can produce enlarged and bizarrelyshaped myocytes as well as disorientation (oblique orperpendicular alignment) of adjacent cells (Figure 2B).These findings, collectively termed myocyte disarray,can be focal and juxtaposed beside normal appearingmyocardium or can be widespread throughout the ven-tricle. With premature death of hypertrophic myocytes,the cardiac fibroblasts and associated extracellular ma-trix increase (often referred to as “replacement” fibrosis)Figure 1. Patterns of Cardiac Remodelingand further contribute to the distortion in myocardialThe normal myocardium can be remodeled by heritable gene muta-

tions and acquired diseases into hypertrophic or dilated morpholo- cell architecture.gies. Note the normal left ventricular wall thickness and chamber Despite the presence of even markedly abnormalvolume of the healthy heart (B) in comparison to the robust increase ventricular morphology and histopathology, contractilein left ventricular wall thickness of hypertrophic cardiomyopathy (A) (systolic) function in hypertrophic cardiomyopathy isand the marked increase in left ventricular chamber volumes of

usually excellent and can often appear supra normal.dilated cardiomyopathy (C). Atrial enlargement is also evident inYet, most affected individuals develop mild to moderateboth cardiomyopathies.symptoms of shortness of breath (dypsnea) and chestpain (angina) due to impaired diastolic relaxation of the

diomyopathy have been defined, genetic mapping stud- hypertrophic heart (Spirito et al., 1997). In addition, pa-ies indicate this disorder will similarly exhibit substantial tients are at risk for heart failure, atrial and ventricularinter- and intragenic heterogeneity. Restrictive cardio- arrhythmias, and sudden death. In the United States,myopathy remains an enigma; neither disease loci nor unrecognized hypertrophic cardiomyopathy is the mostmutated genes have been discovered to cause this rarer common cause for sudden death in athletes.type of cardiomyopathy. Diagnosis of hypertrophic cardiomyopathy is made

The importance of identifying precise genetic causes by two-dimensional echocardiography, a noninvasivefor cardiomyopathies may extend beyond the molecular cardiac imaging technique that reveals the extent andinsights into these specific maladies. With the recent distribution of hypertrophy and delineates cardiac con-development and analyses of genetically engineered tractile function. Echocardiograms from a large popula-models of human cardiomyopathy mutations comes the tion of young individuals have estimated the incidence ofopportunity to define critical molecules and pathways hypertrophic cardiomyopathy to be 1 in 500 individualsthat participate in cardiac remodeling. Since similar pat- (Maron et al., 1995). The wide-spread use of echocardi-terns of cardiac remodeling occur in response to more ography has not only confirmed autosomal dominantprevalent, acquired cardiovascular diseases, mechanis- transmission of disease, it has identified age-dependenttic insights derived from the study of genetic disorders penetrance of the disease and documented marked vari-may be relevant to a wide range of heart conditions that ation in the extent and distribution of the hypertrophy.remodel the heart and contribute to heart failure. Molecular genetic studies of familial hypertrophic car-

diomyopathy help to explain the clinical diversity of thisHypertrophic Cardiomyopathy condition. Linkage analyses provided the first evidenceA primary disorder of the myocardium, hypertrophic car- for allelic heterogeneity with sequential definition of dis-diomyopathy causes distinctive anatomic and histologic ease loci on chromosomes 14q11 (Jarcho et al., 1989),features as well as a wide array of clinical manifestations 1q32 (Watkins et al., 1993), 15q22 (Thierfelder et al.,(Fatkin et al., 2000a and references therein). Cardiac 1993), and 11p11 (Carrier et al., 1993). Demonstrationmass is increased due to left ventricular wall thickening that human mutations in the � cardiac myosin heavy(hypertrophy) that most often is asymmetric, often with chain, cardiac troponin T, � tropomyosin, and cardiacparticular involvement of the interventricular septum. myosin binding protein C genes were encoded at theseAs a consequence of hypertrophy, the left ventricular loci, led to the conclusion that hypertrophic cardiomyop-chamber volumes are diminished, resulting in the ap- athy resulted from defects in cardiac sarcomere proteinspearance of a muscle-bound heart (Figure 1A). Histopa- (for a review, see Seidman and Seidman, 2000). Muta-thology in hypertrophic cardiomyopathy can be strik- tions in cardiac actin (Mogensen et al., 1999), troponin I

(Kimura et al., 1997), the essential and regulatory myosiningly abnormal. The highly registered alignment of

Figure 2. Histopathology of Hypertrophic andDilated Cardiomyopathy

(A) The normal architecture of healthy ventric-ular myocardium shows orderly alignment ofmyocytes with minimal interstitial fibrosis.(B) Marked enlargement and disarray of myo-cytes (red) with increased interstitial fibrosis(blue) is evident in hypertrophic cardiomy-opathy.

(C) The histology of dilated cardiomyopathy shows hypertrophy and degeneration of myocytes (dark red) without disarray. Increases ininterstitial fibrosis (pale pink) is evident. (Stains: [A and C], hematoxylin and eosin; [B], mason trichrome)

Normal Diseased

Cell558

myocytes that characterize the normal myocardium (Fig-ure 2A) becomes distorted by hypertrophic growth ofthe myocytes, which can produce enlarged and bizarrelyshaped myocytes as well as disorientation (oblique orperpendicular alignment) of adjacent cells (Figure 2B).These findings, collectively termed myocyte disarray,can be focal and juxtaposed beside normal appearingmyocardium or can be widespread throughout the ven-tricle. With premature death of hypertrophic myocytes,the cardiac fibroblasts and associated extracellular ma-trix increase (often referred to as “replacement” fibrosis)Figure 1. Patterns of Cardiac Remodelingand further contribute to the distortion in myocardialThe normal myocardium can be remodeled by heritable gene muta-

tions and acquired diseases into hypertrophic or dilated morpholo- cell architecture.gies. Note the normal left ventricular wall thickness and chamber Despite the presence of even markedly abnormalvolume of the healthy heart (B) in comparison to the robust increase ventricular morphology and histopathology, contractilein left ventricular wall thickness of hypertrophic cardiomyopathy (A) (systolic) function in hypertrophic cardiomyopathy isand the marked increase in left ventricular chamber volumes of

usually excellent and can often appear supra normal.dilated cardiomyopathy (C). Atrial enlargement is also evident inYet, most affected individuals develop mild to moderateboth cardiomyopathies.symptoms of shortness of breath (dypsnea) and chestpain (angina) due to impaired diastolic relaxation of the

diomyopathy have been defined, genetic mapping stud- hypertrophic heart (Spirito et al., 1997). In addition, pa-ies indicate this disorder will similarly exhibit substantial tients are at risk for heart failure, atrial and ventricularinter- and intragenic heterogeneity. Restrictive cardio- arrhythmias, and sudden death. In the United States,myopathy remains an enigma; neither disease loci nor unrecognized hypertrophic cardiomyopathy is the mostmutated genes have been discovered to cause this rarer common cause for sudden death in athletes.type of cardiomyopathy. Diagnosis of hypertrophic cardiomyopathy is made

The importance of identifying precise genetic causes by two-dimensional echocardiography, a noninvasivefor cardiomyopathies may extend beyond the molecular cardiac imaging technique that reveals the extent andinsights into these specific maladies. With the recent distribution of hypertrophy and delineates cardiac con-development and analyses of genetically engineered tractile function. Echocardiograms from a large popula-models of human cardiomyopathy mutations comes the tion of young individuals have estimated the incidence ofopportunity to define critical molecules and pathways hypertrophic cardiomyopathy to be 1 in 500 individualsthat participate in cardiac remodeling. Since similar pat- (Maron et al., 1995). The wide-spread use of echocardi-terns of cardiac remodeling occur in response to more ography has not only confirmed autosomal dominantprevalent, acquired cardiovascular diseases, mechanis- transmission of disease, it has identified age-dependenttic insights derived from the study of genetic disorders penetrance of the disease and documented marked vari-may be relevant to a wide range of heart conditions that ation in the extent and distribution of the hypertrophy.remodel the heart and contribute to heart failure. Molecular genetic studies of familial hypertrophic car-

diomyopathy help to explain the clinical diversity of thisHypertrophic Cardiomyopathy condition. Linkage analyses provided the first evidenceA primary disorder of the myocardium, hypertrophic car- for allelic heterogeneity with sequential definition of dis-diomyopathy causes distinctive anatomic and histologic ease loci on chromosomes 14q11 (Jarcho et al., 1989),features as well as a wide array of clinical manifestations 1q32 (Watkins et al., 1993), 15q22 (Thierfelder et al.,(Fatkin et al., 2000a and references therein). Cardiac 1993), and 11p11 (Carrier et al., 1993). Demonstrationmass is increased due to left ventricular wall thickening that human mutations in the � cardiac myosin heavy(hypertrophy) that most often is asymmetric, often with chain, cardiac troponin T, � tropomyosin, and cardiacparticular involvement of the interventricular septum. myosin binding protein C genes were encoded at theseAs a consequence of hypertrophy, the left ventricular loci, led to the conclusion that hypertrophic cardiomyop-chamber volumes are diminished, resulting in the ap- athy resulted from defects in cardiac sarcomere proteinspearance of a muscle-bound heart (Figure 1A). Histopa- (for a review, see Seidman and Seidman, 2000). Muta-thology in hypertrophic cardiomyopathy can be strik- tions in cardiac actin (Mogensen et al., 1999), troponin I

(Kimura et al., 1997), the essential and regulatory myosiningly abnormal. The highly registered alignment of

Figure 2. Histopathology of Hypertrophic andDilated Cardiomyopathy

(A) The normal architecture of healthy ventric-ular myocardium shows orderly alignment ofmyocytes with minimal interstitial fibrosis.(B) Marked enlargement and disarray of myo-cytes (red) with increased interstitial fibrosis(blue) is evident in hypertrophic cardiomy-opathy.

(C) The histology of dilated cardiomyopathy shows hypertrophy and degeneration of myocytes (dark red) without disarray. Increases ininterstitial fibrosis (pale pink) is evident. (Stains: [A and C], hematoxylin and eosin; [B], mason trichrome)

Cell558

myocytes that characterize the normal myocardium (Fig-ure 2A) becomes distorted by hypertrophic growth ofthe myocytes, which can produce enlarged and bizarrelyshaped myocytes as well as disorientation (oblique orperpendicular alignment) of adjacent cells (Figure 2B).These findings, collectively termed myocyte disarray,can be focal and juxtaposed beside normal appearingmyocardium or can be widespread throughout the ven-tricle. With premature death of hypertrophic myocytes,the cardiac fibroblasts and associated extracellular ma-trix increase (often referred to as “replacement” fibrosis)Figure 1. Patterns of Cardiac Remodelingand further contribute to the distortion in myocardialThe normal myocardium can be remodeled by heritable gene muta-

tions and acquired diseases into hypertrophic or dilated morpholo- cell architecture.gies. Note the normal left ventricular wall thickness and chamber Despite the presence of even markedly abnormalvolume of the healthy heart (B) in comparison to the robust increase ventricular morphology and histopathology, contractilein left ventricular wall thickness of hypertrophic cardiomyopathy (A) (systolic) function in hypertrophic cardiomyopathy isand the marked increase in left ventricular chamber volumes of

usually excellent and can often appear supra normal.dilated cardiomyopathy (C). Atrial enlargement is also evident inYet, most affected individuals develop mild to moderateboth cardiomyopathies.symptoms of shortness of breath (dypsnea) and chestpain (angina) due to impaired diastolic relaxation of the

diomyopathy have been defined, genetic mapping stud- hypertrophic heart (Spirito et al., 1997). In addition, pa-ies indicate this disorder will similarly exhibit substantial tients are at risk for heart failure, atrial and ventricularinter- and intragenic heterogeneity. Restrictive cardio- arrhythmias, and sudden death. In the United States,myopathy remains an enigma; neither disease loci nor unrecognized hypertrophic cardiomyopathy is the mostmutated genes have been discovered to cause this rarer common cause for sudden death in athletes.type of cardiomyopathy. Diagnosis of hypertrophic cardiomyopathy is made

The importance of identifying precise genetic causes by two-dimensional echocardiography, a noninvasivefor cardiomyopathies may extend beyond the molecular cardiac imaging technique that reveals the extent andinsights into these specific maladies. With the recent distribution of hypertrophy and delineates cardiac con-development and analyses of genetically engineered tractile function. Echocardiograms from a large popula-models of human cardiomyopathy mutations comes the tion of young individuals have estimated the incidence ofopportunity to define critical molecules and pathways hypertrophic cardiomyopathy to be 1 in 500 individualsthat participate in cardiac remodeling. Since similar pat- (Maron et al., 1995). The wide-spread use of echocardi-terns of cardiac remodeling occur in response to more ography has not only confirmed autosomal dominantprevalent, acquired cardiovascular diseases, mechanis- transmission of disease, it has identified age-dependenttic insights derived from the study of genetic disorders penetrance of the disease and documented marked vari-may be relevant to a wide range of heart conditions that ation in the extent and distribution of the hypertrophy.remodel the heart and contribute to heart failure. Molecular genetic studies of familial hypertrophic car-

diomyopathy help to explain the clinical diversity of thisHypertrophic Cardiomyopathy condition. Linkage analyses provided the first evidenceA primary disorder of the myocardium, hypertrophic car- for allelic heterogeneity with sequential definition of dis-diomyopathy causes distinctive anatomic and histologic ease loci on chromosomes 14q11 (Jarcho et al., 1989),features as well as a wide array of clinical manifestations 1q32 (Watkins et al., 1993), 15q22 (Thierfelder et al.,(Fatkin et al., 2000a and references therein). Cardiac 1993), and 11p11 (Carrier et al., 1993). Demonstrationmass is increased due to left ventricular wall thickening that human mutations in the � cardiac myosin heavy(hypertrophy) that most often is asymmetric, often with chain, cardiac troponin T, � tropomyosin, and cardiacparticular involvement of the interventricular septum. myosin binding protein C genes were encoded at theseAs a consequence of hypertrophy, the left ventricular loci, led to the conclusion that hypertrophic cardiomyop-chamber volumes are diminished, resulting in the ap- athy resulted from defects in cardiac sarcomere proteinspearance of a muscle-bound heart (Figure 1A). Histopa- (for a review, see Seidman and Seidman, 2000). Muta-thology in hypertrophic cardiomyopathy can be strik- tions in cardiac actin (Mogensen et al., 1999), troponin I

(Kimura et al., 1997), the essential and regulatory myosiningly abnormal. The highly registered alignment of

Figure 2. Histopathology of Hypertrophic andDilated Cardiomyopathy

(A) The normal architecture of healthy ventric-ular myocardium shows orderly alignment ofmyocytes with minimal interstitial fibrosis.(B) Marked enlargement and disarray of myo-cytes (red) with increased interstitial fibrosis(blue) is evident in hypertrophic cardiomy-opathy.

(C) The histology of dilated cardiomyopathy shows hypertrophy and degeneration of myocytes (dark red) without disarray. Increases ininterstitial fibrosis (pale pink) is evident. (Stains: [A and C], hematoxylin and eosin; [B], mason trichrome)

Shown in this example is myosin of that powers the contraction of muscle cells. The colored dots show individual amino acids which when changed alter the activity of myosin. We can correlate the changes in myosin sequence and activity with changes in the structure and function of cells and tissues. In this case, cardiac muscle cells that mediate contraction of the heart. The changes in cell function can be seen to alter the overall structure of organs, not the increased size of the muscle tissue in the heart with the mutation in muscle myosin and the decreased volume of the left ventricle. Finally, the changes in the structure of the heart compromise its activity leading to disease and often death. So how do we think about cells?

We use models to explain biological events.

DNA

RNA

Protein

To help us understand the events that take place in cells we use models to represent the processes and pathways that have been discovered experimentally. Some models are simple, have lots of supporting data and and appear elegant, and some might say beautiful. However, these models usually hide the underlying details that make the events more complicated and less elegant. Ugly? Perhaps, but it’s a more accurate reflection of reality.

No one person can understand these processes and the connections between these events and medicine and disease are often tenuous and unclear. How can we make sense of all these molecular events so that they generate helpful information for biology and medicine?

We use models to explain biological events.

To help us understand the events that take place in cells we use models to represent the processes and pathways that have been discovered experimentally. Some models are simple, have lots of supporting data and and appear elegant, and some might say beautiful. However, these models usually hide the underlying details that make the events more complicated and less elegant. Ugly? Perhaps, but it’s a more accurate reflection of reality.

No one person can understand these processes and the connections between these events and medicine and disease are often tenuous and unclear. How can we make sense of all these molecular events so that they generate helpful information for biology and medicine?

We use models to explain biological events.

To help us understand the events that take place in cells we use models to represent the processes and pathways that have been discovered experimentally. Some models are simple, have lots of supporting data and and appear elegant, and some might say beautiful. However, these models usually hide the underlying details that make the events more complicated and less elegant. Ugly? Perhaps, but it’s a more accurate reflection of reality.

No one person can understand these processes and the connections between these events and medicine and disease are often tenuous and unclear. How can we make sense of all these molecular events so that they generate helpful information for biology and medicine?

Cellular events can be organizes into systems that generate specific outcomes and products.

Metabolism

Structure

Communication

Organization

Adhesion

Growth/Division

Secretion

One proposed approach is called systems cell biology that suggests that a cell consist of a set of systems that contribute to its viability and function. Each system generates unique outcomes or products that contribute to the health of a cell. Each system employs thousands of molecules to generate its outcomes and products but instead of focusing on individual molecules, we look at the overall structure and performance of each system. Each system is capable of autoregulation, can regulate its activity based on environmental conditions. Systems are dependent on each other.

This course will explore each of these systems and more. Some are covered in more detail in other courses (metabolism). Will discuss the role of each system in cell function and viability and how changes in each system lead to disease. Where the course presents molecular details, it does so for several reasons. First, the details may be linked to certain diseases, so a mutation in a specific protein may lead to a specific disease. The details are a mechanism common to different systems. Finally, the details are cool.

Plasma membrane: defining the boundary of cell

Plasma membrane defines the outer limit of cells.

The plasma membrane defines the border of a cell. It prevents the loss of cellular material and restricts the passage of material from the external environment. Why is it needed? Other objects stay together without membranes. Proteins and other cellular material undergo rapid diffusion due to thermal energy. Something must prevent material from diffusing away from cell.

Membranes are selectively permeable.

Proteins

NucleotidesAmino AcidsSugars

Na+, K+, Ca2+, Cl-, H+, Mg2+

O2, CO2, NOSteroids

1. Membranes restrict passage based on size and charge.2. Proteins can’t diffuse across membranes and most large molecules (DNA, RNA, carbohydrates) can’t move freely across membranes.3. Even smaller subunits of proteins, nucleic acids and carbohydrates are too large to pass.4. Ions can’t pass.

1. Very small but charge prevents their passage across membrane.5. Gases freely diffuse.

1. Exchange oxygen and CO2. NO a key signaling molecule.6. Steroids diffuse.

1. Larger but hydrophobic.2. Signaling molecules.

Membranes contain hydrophilic outer surfaces and a hydrophobic core.

What makes membranes an effective barrier to prevent diffusion of proteins and ions is their structure. Membranes consist of a bilayer of phospholipids. The layers of phospholipids are arranged into an outer leaflet that faces the external environment and an inner leaflet that faces the inside of the cell. Membranes have two key chemical properties. The outer surfaces is charged and is soluble in water. All cells exist in aqueous environments. The central core of membranes, in contrast, is hydrophobic or repels water. The hydrophobic core is what prevents the diffusion of charge molecules and ions. The ability of phospholipids to pack together tightly is what restricts the diffusion of larger molecules.

Phospholipids contain a hydrophilic head group and hydrophobic tail.

The cartoon illustrates the general structure of a phospholipid that composes membranes. There are two important chemical and structural features of phospholipids. The polar head group makes phospholipids soluble in water. The long hydrophobic tails allows phospholipids to self-assemble into bilayers to form membranes. The hydrophobic tails are composed of long chains of hydrocarbons. If the carbons are all linked by single bonds, then the tail is called saturated and tends to be more straight. Saturated lipids pack more closely together. A double-bond introduces a kink in the tail and prevents lipids from packing as closely together as in saturated lipids. Lipids with a double bond are called unsaturated. Biological membranes contain a mix of saturated and unsaturated lipids as a membrane with all saturated lipids would be a solid at physiological temperature.

Phosphoglycerides can have a variety of head groups.

Membranes are comprised of a variety of phospholipids and other lipids. The major difference between phospholipids is in the chemical composition of the polar head group. This slide illustrated the different types of lipids in a class called phosphoglycerides which compose the major type of lipid in membrane. There are several different polar head groups with different shapes and charges. Proteins can distinguish between the different head groups and cells can recruit specific proteins to membranes by adjusting the composition of phosphoglycerides.Another type of phosphoglycerides is inositols that are critical in cell signaling.

Plasma membrane allows for ion gradients between cytosol and extracellular fluid.

[Na+] ~ 15 mM

[Na+] ~ 145 mM

[K+] ~ 120 mM

[K+] ~ 4.5 mM

-70 mV

As I mentioned, membranes restrict the diffusion of ions and this has important biological consequences. One is that it allows cells to create ion gradients. For most cells, the concentration of sodium is significantly higher outside the cell than inside. For potassium, the reverse is true: the concentration inside the cell is higher than outside the cell. The distribution of other ions also differs between inside and outside. One consequence of this asymmetric distribution of ions is that the overall charge inside the cell is negative compared to the outside. This electrical potential across the membrane is critical for cell communication, in particular between neurons where action potentials that travel down axons proceed by lowering this potential difference across the membrane.

Membranes keep cytosolic calcium low and enzymes inactive.

[Ca2+] ~ .0001 mM

[Ca2+] ~ 1.2 mM

[Ca2+] ~ .3 mM

The other important ion that membranes restrict is calcium. Cells keep the cytosolic concentration of calcium very low because calcium activates a variety of cellular enzymes. When cells need to activate those enzymes, they increase the concentration of cytosolic calcium. Cells intensively regulate cytosolic calcium because prolonged activation of some enzymes can lead to cell damage and eventually cell death.

Damage to plasma membrane increases cytosolic calcium and can lead to cell death.

[Ca2+] ~ .0001 mM

[Ca2+] ~ 1.2 mM

[Ca2+] ~ .3 mM

Importantly, any damage to the plasma membrane allows calcium to enter the cell and activate several cellular enzymes. Consequently, cells have active processes involving the secretory pathway that allows them to repair damage to the plasma membrane. Cells that are exposed to mechanical stress, muscle and cells that line blood vessels, are especially susceptible to damage of their plasma membranes. Muscular dystrophy which is the weakening of skeletal muscle results from excessive damage to the plasma membrane of skeletal muscle cells. These cells are not able to keep up with the repair of their plasma membranes and eventually die.

Damage to plasma membrane increases cytosolic calcium and can lead to cell death.

[Ca2+] ~ .0001 mM

[Ca2+] ~ 1.2 mM

[Ca2+] ~ .3 mM

Importantly, any damage to the plasma membrane allows calcium to enter the cell and activate several cellular enzymes. Consequently, cells have active processes involving the secretory pathway that allows them to repair damage to the plasma membrane. Cells that are exposed to mechanical stress, muscle and cells that line blood vessels, are especially susceptible to damage of their plasma membranes. Muscular dystrophy which is the weakening of skeletal muscle results from excessive damage to the plasma membrane of skeletal muscle cells. These cells are not able to keep up with the repair of their plasma membranes and eventually die.

Damage to plasma membrane increases cytosolic calcium and can lead to cell death.

[Ca2+] ~ .0001 mM

[Ca2+] ~ 1.2 mM

[Ca2+] ~ .3 mM

Importantly, any damage to the plasma membrane allows calcium to enter the cell and activate several cellular enzymes. Consequently, cells have active processes involving the secretory pathway that allows them to repair damage to the plasma membrane. Cells that are exposed to mechanical stress, muscle and cells that line blood vessels, are especially susceptible to damage of their plasma membranes. Muscular dystrophy which is the weakening of skeletal muscle results from excessive damage to the plasma membrane of skeletal muscle cells. These cells are not able to keep up with the repair of their plasma membranes and eventually die.

How do cells get essential nutrients and material across plasma membrane?

Proteins

NucleotidesAmino AcidsSugars

Na+, K+, Ca2+, Cl-, H+, Mg2+

O2, CO2, NOSteroids

The plasma membrane restricts the diffusion of a lot of different types of molecules and ions, but cells need theses molecules to grow and survive. How do cells take up this material from the external environment?

Protein channels allow passage of small molecules and ions across membranes.

Sugars Amino acids Ions

Inside

Outside

Plasma membrane is associated with many different types of proteins. One class are channels that span both bilayers and contain a pore that allows that passage of specific molecule or ion. The opening of these pores is tightly regulated and occasionally requires energy.

Receptors allow cells to sense and communicate with outside world.

Inside

Outside

A second type of protein found in the plasma membrane is receptors that allow cells to sense and respond to the external environment. Receptors interact with specific molecules and chemicals and relay their binding state across the membrane to activate or inactivate different cellular events.

Proteins and lipids diffuse rapidly in membranes.

1. One other critical point is that lipids are not static within a membrane. They show a great deal of movement.2. The tails of lipids can bend or rotate due to thermal energy.3. And very rarely, a lipid will flip from one leaflet to another.4. But most important is that lipids diffuse within membranes.

4.1. Shown here experimentally. 4.2. Label lipids with fluorescent dye.4.3. Use laser to bleach dye in certain area.4.4. Measure recovery of fluorescence in bleached area -> fluorescence lipids can only come from surrounding areas.4.5. If fluorescence returns, lipids must be able to move laterally.4.6. Rate of recovery gives estimate of how fast lipids move laterally in membranes.4.7. 1 µm2/s -> circumnavigate bacteria in few seconds.

Membranes are fluid.

Cytoskeleton provides structural support for plasma membrane.

Plasma membrane

CytoskeletonMembrane proteins

The cytoskeleton provides mechanical support to the plasma membrane. Actin filaments are the primary cytoskeletal filaments that support the plasma membrane. Actin filaments can be arranged in a variety of configurations to create different shapes of the plasma membrane (see lecture on cell shape for details). In this image, the filaments are arranged in a mesh and tethered to the plasma membrane by interactions with integral membrane proteins. This arrangement of actin filaments provides mechanical support to a large area of the plasma membrane.