ABSTRACT

TRIM46 is a RING finger E3 ligase which belongs to TRIM (tripartite motif-containing) protein family. TRIM46 is required for neuronal polarity and axon specification by driving the formation of parallel microtubule arrays, whereas its embryological functions remain to be determined yet. Expression patterns and biological functions of trim46a, a zebrafish homologue of TRIM46, were studied in zebrafish embryo. First, maternal transcripts of trim46a were present at 1 cell stage whereas zygotic messages were abundant in the eyes, MHB (Midbrain-Hindbrain Boundary) and hindbrain at 24 hpf (hours post fertilization). Second, transcriptional regulatory region of trim46a contains cis-acting elements binding a transcriptional factor Foxa2. Transcription of foxa2 is positively regulated by Sonic Hedgehog (SHH), and treatment of cyclopamine, an SHH inhibitor, represses transcription of foxa2 in 4 hpf through 24 hpf embryos. Third, the transcriptional repression of foxa2 inhibited transcription of trim46a to cause developmental defects in the midbrain and MHB. Finally, spatiotemporal expression patterns of a midbrain marker otx2b in the developmental defects confirmed inhibition of SHH by cyclopamine caused underdevelopment of the midbrain and MHB at 24 hpf. We propose a signaling network where trim46a contributes to development of the midbrain and MHB via Foxa2, a downstream element of SHH signaling in zebrafish embryogenesis.

摘要

TRIM46是属于TRIM（含三方基序）蛋白家族的RING手指E3连接酶。通过驱动平行微管阵列的形成，TRIM46是神经元极性和轴突规范所必需的，而其胚胎学功能仍有待确定。研究了TRIM46的斑马鱼同源物trim46a在斑马鱼胚胎中的表达模式和生物学功能。首先，trim46a的母本转录本出现在1个细胞阶段，而合子的信息在24 hpf（受精后数小时）的眼睛，MHB（Midbrain-Hindbrain边界）和后脑中很丰富。第二，trim46a的转录调控区含有顺式-作用元件结合转录因子Foxa2。foxa2的转录受到Sonic Hedgehog（SHH）的正调控，对SHH抑制剂环巴胺的治疗可抑制foxa2在4 hpf至24 hpf胚胎中的转录。三，转录抑制FOXA2的抑制转录trim46a引起脑和MHB发育缺陷。最后，发育缺陷中的中脑标志物otx2b的时空表达模式证实了环丙胺对SHH的抑制作用在24 hpf时引起中脑和MHB的发育不足。我们提出了一个信号网络，其中trim46a 通过Foxa2（斑马鱼胚胎发生中SHH信号的下游元素）促进中脑和MHB的发育。