Analysis of the cell adhesion molecule sticks-and-stones reveals multiple redundant functional domains, protein-interaction motifs and phosphorylated tyrosines that direct myoblast fusion in Drosophila melanogaster.
The larval body wall muscles of Drosophila melanogaster arise by fusion of founder myoblasts (FMs) and fusion-competent myoblasts (FCMs). Sticks-and-Stones (SNS) is expressed on the surface of all FCMs and mediates adhesion with FMs and developing syncytia. Intracellular components essential for myoblast fusion are then recruited to these adhesive contacts. In the studies herein, a functional analysis of the SNS cytodomain using the GAL4/UAS system identified sequences that direct myoblast fusion, presumably through recruitment of these intracellular components. An extensive series of deletion and site-directed mutations were evaluated for their ability to rescue the myoblast fusion defects of sns mutant embryos. Deletion studies revealed redundant functional domains within SNS. Surprisingly, highly conserved consensus sites for binding post-synaptic density-95/discs large/zonula occludens-1-domain-containing (PDZ) proteins and serines with a high probability of phosphorylation play no significant role in myoblast fusion. Biochemical studies establish that the SNS cytodomain is phosphorylated at multiple tyrosines and their site-directed mutagenesis compromises the ability of the corresponding transgenes to rescue myoblast fusion. Similar mutagenesis revealed a requirement for conserved proline-rich regions. This complexity and redundancy of multiple critical sequences within the SNS cytodomain suggest that it functions through a complex array of interactions that likely includes both phosphotyrosine-binding and SH3-domain-containing proteins.
Kocherlakota
Kiranmai S
KS
Stowers Institute for Meical Research, Kansas City, Missouri 64110, USA.
Wu
Jian-Min
JM
McDermott
Jeffrey
J
Abmayr
Susan M
SM
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D015815:Cell Adhesion Molecules / Q000737:chemistry*; D002459:Cell Fusion; D017124:Conserved Sequence; D029721:Drosophila Proteins / Q000737:chemistry*; D004331:Drosophila melanogaster / Q000166:cytology* / Q000196:embryology; D004625:Embryo, Nonmammalian / Q000166:cytology; D007136:Immunoglobulins / Q000737:chemistry*; D008969:Molecular Sequence Data; D032446:Myoblasts / Q000166:cytology*; D010641:Phenotype; D010768:Phosphoserine / Q000378:metabolism; D019000:Phosphotyrosine / Q000378:metabolism*; D011392:Proline; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D019076:Transgenes
D015815:Cell Adhesion Molecules; D029721:Drosophila Proteins; D007136:Immunoglobulins; C410904:SNS protein, Drosophila; D010768:Phosphoserine; D019000:Phosphotyrosine; D011392:Proline