olga gonzalez1
TRANSCRIPT
Herpesviral reactivation: A key step for disease development.
Olga D. Gonzalez-Lopez, Ph.D candidate University of Medicine and Dentistry of New Jersey
Dept. Microbiology and Molecular Genetics
• Introduce myself
• Overview of the Human Herpesvirus family
• Molecular mechanism of reactivation in Kaposi’s sarcoma-associated herpesvirus (KSHV).
– Promoter specification by Rta and RBP-Jk
• GSBS Summer Research Program @ UMDNJ
Activation of lytic program
Establish Latency
Maintain Latency
Reactivation
Latency:
life-long persistent infection. poor target of host immune response. enhances cell survival and promotes proliferation. proper segregation of the viral episomes into daughter cells during mitosis.
Reactivation
α β
γ
HSV-1
VZV
CMV HHV-6
HHV-7
EBV
KSHV
HSV-2
Kimberlin, DW. 2005. Semin Pediatr Infect Dis. 16(4): 271–281.
HSV-1: infections above the waist
HSV-2: infections below the waist
Diseases:
o Oral herpes or cold sore
o Genital herpes
Treatment: Acyclovir
Treatment: Live attenuated vaccine
Congenital disease:
o Virus can spread via the placenta to the fetus and congenital abnormalities can occur
o Different route of transmission: breast milk, during birth
Centers for Disease Control and Prevention (CDC): h"p://www.cdc.gov/features/dscytomegalovirus/
Treatment: Ganciclovir
Epstein Barr Virus Kaposi’s sarcoma-associated herpesvirus
95% of people in the U.S. have been infected with EBV.
Up to 80% of students entering college in the US are seropositive.
Infectious mononucleosis
Burkitt lymphoma Nasopharyngeal carcinoma
NO TREATMENT
J. Bras. Pneumol. vol.31 no.6 São Paulo Nov./Dec. 2005
h"p://www.dermis.net/dermisroot/en/17372/image.htm
TPA
HDAC inhibitors
IE genes DE genes Late genes vDNA-R
TFs IE
Rta
vector!
ORF50/Rta! TPA!
untreated!
TranscripIonal AcIvaIon Domain
DNA binding
NLS=Nuclear localiza/on signal
TetramerizaIon
Rta
Notch signaling pathway is evolutionarily conserved pathway regulate cell fate decision in various cell types during and after development
Aberrant Notch signaling tumor formation and progression.
1
(45 kb.)
(90 kb.)
(140 kb.)
3x 2x 3x 2x 2x
Jk x
Only Rta, but not the RBP-Jk-dependent activators NICD-1 nor EBV EBNA2, productively reactivate KSHV from latency
RBP‐Jk is not consItuIvely and broadly bound to KSHV DNA.
CANT CANT CANT CANT
Rta Rta Rta
RBP-Jk Rta sImulates DNA binding of RBP‐Jk.
Rta
Rta transacIvaIon
308.0x (19.0) TATA ‐30
‐957 Luciferase Jk ‐136 ‐106 ‐54
7.6x (0.6) TATA ‐30
‐957 Luciferase Jk ‐136
X ‐106 ‐54
HDAC
X X X X Ac Ac Ac Ac Ac
RBP- Jk VP16 AD
X RBP-Jk
HDAC
RBP-Jk
RBP- Jk VP16 AD
1 530 RtaΔSTAD
Rta WT
TATA ‐30
‐957 Luciferase Jk ‐136 ‐106 ‐54
Mta Promoter: ‐957 ‐136
[A/T]3 - N7 - [A/T]3 N7 - [A/T]3
[A/T]3 - N17 - [A/T]3
Liao, et al., 2003
Ziegelbauer, et al., 2006
CCCACTTC
Palmeri, et al., 2011
ANTGTAACANTA/TA/TT
Guito,J and Lukac DM, 2012.
G + A ladder
0 0
0 + + + ++ +
3’AGTGCCATTGTGAATACTCAGTCACAAAACGGTCGTTCACATTGTTATTACAAGGGTGCCGGGTAAAAAGCAAAC 5’ RBP‐Jk
‐136 ‐62 Mta
3’AGTGCCATTGTGAATACTCAGTCACAAAACGGTCGTTCACATTGTTATTACAAGGGTGCCGGGTAAAAAGCAAAC 5’ 5’TCACGGTAACACTTATGAGTCAGTGTTTTGCCAGCAAGTGTAACAATAATGTTCCCACGGCCCATTTTTCGTTTG3’
‐62 ‐136
RBP‐Jk
Palmeri, et al. 2011
3’AGTGCCATTGTGAATACTCAGTCACAAAACGGTCGTTCACATTGTTATTACAAGGGTGCCGGGTAAAAAGCAAAC 5’ 5’TCACGGTAACACTTATGAGTCAGTGTTTTGCCAGCAAGTGTAACAATAATGTTCCCACGGCCCATTTTTCGTTTG 3’
1R 2R 3R
1F 2F 3F 4F
‐62 ‐136
RBP‐Jk
Palmeri, et al. 2011
1F 2F 3F 4F 1R 2R 3R
CONS
A T A C G A G
A
C A G A C T T
N
G T T CCTG
T
GGGGAGG
G
TATGATG
T
AGACATA
A
ATACAAA
A
CCCCCCC
C
AAAAAAA
A
CGATCCT
N
TTTTTTT
T
TGATGTA
ATATAGT
TTTTCCT
T A/TA/T
‐136
0 5 10 15 20 25 30
3R 2F
1R 2R 1F 4F 3F Jk
4F 3R
3F Jk
4F Jk
Jk
4F 3R
3F Jk
3R 2F
1R 2R 3F Jk
3R 2F
1R 3F Jk
E
F
G
H
I
J
Palmeri, et al. 2011
RBP‐Jk
Rta
RBP-Jk
Rta Rta
Rta Rta Rta
Rta Rta Rta
3’AGTGCCATTGTGAATACTCAGTCACAAAACGGTCGTTCACATTGTTATTACAAGGGTGCCGGGTAAAAAGCAAAC 5’ 5’TCACGGTAACACTTATGAGTCAGTGTTTTGCCAGCAAGTGTAACAATAATGTTCCCACGGCCCATTTTTCGTTTG3’
‐62 ‐136
Mta
• Understanding of molecular mechanism for disease development.
• Manipulation of host signaling pathways. “Molecular piracy”
• Understanding of cancer development and progression.
• Many cis-elements contribute to Rta/RBP-Jk transactivation.
• The number and positions of CANT and [A/T]3 repeats and their relative positions to the RBP-Jk binding site, determines Rta transactivation.