Zebrafish Bmal1b Is Essential For Circadian Regulation And Plays A Role In The Reproductive System

The circadian clock exists in almost every kind of living organisms，and plays animportant role in maintaining their normal function. A transcription/translation-basednegative feedback loop composed of a group of circadian clock genes is known toregulate the circadian clock. BMAL and CLOCK as the positive elements formheterodimers that bind to E-box (CANNTG) in the promoter regions of the downstreamgenes including Per and Cry and activate their transcription. Translated protein PER andCRY as negative elements form heterodimers that bind to the CLOCK-BMALheterodimer, turn off their own transcription, thus close the loop. There are two Bmalgenes in mice. Gene targeting studies showed that Bmal1, rather than Bmal2, is essentialfor maintaining the rhythmicity of their gene expression and locomotor activity. Thereare three bmal genes i.e., bmal1a, bmal1b and bmal2, in zebrafish. The circadianfunctions of these three zebrafish bmal genes are not yet investigated.Here, we study the ENU-induced bmal1b mutant and generate the bmal2mutantsusing TALENs system. We found down-regulated gene expression of per1a, per1b,cry1ab and elevated locomotor activity in bmal1b mutant, which are consistent with theresults of clock1a dominant-negative knock-down transgenic zebrafish. We also foundreduced level of estradiol concentration in female bmal1b mutants. q-PCR showeddown-regulations of StAR and cyp19a1a in female bmal1b mutants. These studiesshowed bmal1b is essential for circadian regulation and plays a role in the reproductivesystem.In addition, three types of bmal2homozygous mutants also were generated usingthe TALENs system, which should set the stage for studying its circadian functions inthe future.