Part I characterization of MyoR in C2C12 mouse fibroblasts. Part II isolation and characterization of a novel class II bHLH transcription factor from the black widow spider, latrodectus hesperus

• Main Author: Thayer, William R.
• Format: Others
• Published: Scholarly Commons 2004
• Subjects: Transcription factors; Fibroblasts; Mice Physiology; Black widow spider Physiology; Life Sciences
• Online Access: https://scholarlycommons.pacific.edu/uop_etds/598; https://scholarlycommons.pacific.edu/cgi/viewcontent.cgi?article=1597&context=uop_etds

PART I The basic helix-loop-helix (bHLH) family of transcription factors are involved in a variety of developmental processes. MyoR is the mouse homologue of the human transcription factor ABF-1 . MyoR is classified as a class II basic-helix-loop-helix transcription factor. In order to better understand the relationship between MyoR and muscle cell differentiation, we analyzed the temporal expression at both the mRNA and protein level. Unlike previous studies, we have utilized reverse transcriptase quantitative PCR to analyze mRNA expression. This allows quantitative analysis of MyoR mRNA levels during muscle cell differentiation. We have also analyzed MyoR expression at the protein level. Our studies suggest that the temporal expression of MyoR at the mRNA level is similar to the expression profile seen at the protein level. To ascertain differences in the MyoR DNA-binding activity during myogenesis we performed EMSA. Results suggest that changes in MyoR expression fail to account for differences in the DNAbinding complexes to an E-box site. Part II Members of the basic helix-loop-helix (bHLH) family are required for a number of different developmental pathways, including lymphopoiesis, myogenesis, neurogenesis and sex determination. Screening a eDNA library prepared from silk-producing glands of the black widow spider, we have identified a new bHLH transcription factor named BW6. Within the bHLH region, BW6 shows considerable conservation with other HLH proteins, including Drosophila melanogaster achaete and scute, as well as three HLH proteins identified by gene prediction programs. The expression pattern of bw6 is restricted to a subset of silk producing glands, which includes the tubuliform and major ampullate glands. BW6 is capable of binding an E-box element as a heterodimer with E2A, but was unable to bind this motif as a homodimer. BW6 is also capable of inhibiting the transactivation of rE47 in mammalian cells. BW6 represents the first example of a silk-gland-restricted bHLH protein, and its expression pattern suggests that BW6 may play a role in regulating differentiation of cells in the spider that control silk gland formation or egg case silk gene expression.