Cloning and analysis of small cytoplasmic leucine-rich repeat protein (SCLP), a novel, phylogenetically-conserved protein that is dramatically up-regulated during the programmed death of moth skeletal muscle.

Abstract

We used the abdominal intersegmental muscles (ISMs) of the moth Manduca sexta as a source of transcripts that are dramatically up-regulated during programmed cell death. One of these transcripts, Small Cytoplasmic Leucine-Rich Repeat Protein (SCLP), encodes a protein of approximately 24 kD that contains four perfect and two imperfect leucine-rich repeat (LRR) motifs. DNA sequence database analysis suggests that SCLP is a phylogenetically-conserved gene of unknown function. Both Northern and Western blots demonstrated that SCLP is expressed in the ISMs at all stages examined, but increases greater than 10-fold when the cells become committed to die. This increase in expression is regulated by the same change in the circulating ecdysteroid titer that controls death. Low levels of SCLP expression are also seen in flight muscle and fat body, but not in ovary, male sexual accessory gland, or Malpighian tubules. Immunohistochemical analysis demonstrates that SCLP is a cytoplasmic protein. Western blot analysis of proteins from the fly Drosophila suggests that an SCLP-related protein is expressed at the larval and pupal stages, but not in embryos or adults. Targeted expression of moth SCLP to a variety of different tissues in Drosophila using the Gal4/UAS P element system failed to generate an overt phenotype. These data are interpreted as suggesting that whereas SCLP presumably plays an important role in programmed cell death of muscle, perhaps by acting as an adaptor protein, its expression is insufficient to initiate death by itself.