ANALYSIS OF THE RABBIT CARDIAC SLOW-TWITCH MUSCLE SARCOPLASMIC-RETICULUM CALCIUM-ATPASE (SERCA2) GENE PROMOTER

The rabbit cardiac/slow twitch muscle sarcoplasmic reticulum (SR) Ca2 ATPase (SERCA2) gene encodes a Ca2+ transport pump whose expression i s regulated during skeletal/cardiac muscle development and by differen t pathophysiological states of the heart. This study was designed to d elineate cis-acting regulatory elements involved in SERCA2 gene expres sion. A series of unidirectionally deleted fragments of the upstream 1 ,460 bp SERCA2 promoter were linked to the chloramphenicol acetyltrans ferase (CAT) reporter gene. Transient DNA transfection experiments per formed with these constructs in C2C12 muscle cells and NIH3T3 fibrobla sts revealed a 17 bp upstream promoter element (UPE) important lor tra nscription of the SERCA2 gene in skeletal muscle cells. These studies have also identified a strong (muscle specific) negative regulatory re gion located upstream of nucleotide - 658. Gel mobility shift and sout hwestern analyses using the 17 bp UPE have revealed a specific DNA bin ding complex referred to as Ca2+ ATPase promoter factor -1 (CaPF1). Th e binding factor has an approximate M(r) of 43 kDa. Comparison of CaPF 1 with known transcription factors suggests that the CaPF1 complex may be a novel DNA-binding transcription factor which plays a role in SER CA2 gene regulation in vivo.