Sequence and expression of BET family genes in zebrafish

Drosophila fsh, human RING3, and rat mud6 are homologous members of the BET family, proteins containing dual bromodomains and an extraterminal (ET) domain. Bromodomain proteins are involved in transcriptional regulation; both bromo and ET domains participate in protein–protein interactions. fsh is a maternal effect gene affecting segmentation, RING3 is a mitogenactivated, nuclear kinase that transactivates E2F-dependent genes, and mud6 is upregulated in rat neurons undergoing apoptosis. These data implicate fsh/Ring3/mud6 sequences in early development, cell proliferation, and cell death. To study these sequences in early vertebrate development, the zebrafish homolog (zRing3) was cloned using a probe generated by PCR amplification of the conserved bromodomain. Twelve of the resulting 25 positive clones from a zebrafish embryonic library were sequenced based on unique restriction enzyme digest patterns. Each shows similarity at one or both ends to Ring3. Two categories of Ring3-related cDNAs have been identified. The first contains what appear to be true zRing3 cDNAs, from the structural homolog in zebrafish, and is represented by two size classes of cDNA (zRing3.6 and zRing3.19), encoding proteins that differ only in the N-terminal half of their ET domains. This is the first example of a BET protein with a modified ET domain. The second category contains Ring3-related cDNAs that show sequence similarity only at their 5 ends (zrRing3.9). The chromosomal locations of these cDNAs will be determined to verify the common origin of zRing3.6 and zRing3.19 and to assess whether cDNAs map to regions containing known genetic mutations. The expression pattern of zRing3.6, determined by Northern blot and in situ hybridization, suggests that zRing3 plays a role in the development of head and tail, fin buds, heart, kidney, gut, and regions of the brain. Probes specific to zRing3.6 and zRing3.19 are being used to assess transcript-specific expression patterns during development.