cDNA cloning and tissue-specific expression of a novel basic helix-loop-helix/PAS protein (BMAL1) and identification of alternatively spliced variants with alternative translation initiation site usage

Abstract

Basic helix-loop-helix (bHLH)/PAS proteins, such as Sim, act as transcriptional factors, playing a critical role in the control of central nervous system (CNS) development. To isolate novel bHLH/PAS factors in the CNS an iterative search of a database for expressed sequence tags (ESTs) resulted in the location of several bHLH/PAS protein-like sequences. The rapid amplification of cDNA end (RACE) method was applied to isolate full-length cDNAs of these ESTs. Several 5' and 3' terminal sequences were isolated using primers derived from an EST from the human brain cDNA library. The predicted novel factor polypeptide had bHLH and PAS domains that were highly homologous with those of Ah receptor nuclear translocator (Arnt) and Arnt2. Combination of the isolated cDNA fragments revealed the existence of several alternatively spliced variants. The distribution of the novel bHLH/PAS factor message was analyzed by Northern blot hybridization. This detected only one transcript, which was 2.9 kb in size. Strong hybridization was found in the brain, skeletal muscle and heart. Expression of the novel bHLH/PAS factor, brain and muscle Arnt-like protein 1 (BMAL1), was different from that of Arnt and Arnt2, suggesting that BMAL1 has a different function in the CNS and muscle than Arnt and Arnt2.