molecular characterization of the caenorhabditis elegans alp/enigma gene alp-1 caroline r. mckeown,...
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TRANSCRIPT
Molecular characterization of the Caenorhabditis elegans
ALP/Enigma gene alp-1
Caroline R. McKeown, Hsiao-Fen Han, and Mary C. Beckerle
Developmental Dynamics Vol 235, Issue 2
Muscle Attachment Sites
• Found in dense bodies• Crosslinks actin filaments• Defective -actinin results in
cardiomyopathy (not good)
-Actinin
• ALP (-actinin associated LIM protein)• Family includes at least 7 proteins
• ALP (which has subfamilies including ALP, RIL, CLP36/Elfin, and Mystique)
• Enigma
• Cypher
• All bind either -actinin or -tropomyosin and localize to Z-lines
Mammalian ALP
• LIM (LIN-11, Isl-1, MEC-3) • PDZ (PSD95, Discs-large, ZO-1)
• both are known protein binding interfaces
ALP family domains
C. elegans ALP is eat-1
• First uncovered in a screen by Leon Avery for mutants with pharyngeal pumping defects
• somewhat Unc, also has irregular pharyngeal pumping
• SAGE data shows raised expression levels in larval stages
• similar expression pattern for -actinin
C. elegans ALP is eat-1
C. elegans ALP is eat-1
• Made GFP translational fusions for alp-1
ALP-1 Localization
• All isoforms localize to dense bodies
• ALP-1BCD also found at dense plaques• sites of muscle-muscle adhesion at the muscle termini (vinculin co-localization)
ALP-1 Localization
• Sufficient for localization to dense bodies• not sufficient to localize to dense plaques
• Made PDZ domain containing (and myc-tagged) construct under influence of heat shock promoter
PDZ Domain
• Not essential for viability• PDZ domain localizes protein to sites of
-actinin localization• LIM proteins enable additional protein-
protein interactions• many interactions in Y2H interactome project
• More study needed for specific role
Conclusions
Layton getting CRAZY!!!