Expression pattern and functions of autophagy-related geneatg5in zebrafish organogenesis

Abstract

The implications of autophagy-related genes in serious neural degenerative diseases have been well documented. However, the functions and regulation of the family genes in embryonic development remain to be rigorously studied. Here, we report on for the first time the important role of atg5 gene in zebrafish neurogenesis and organogenesis as evidenced by the spatiotemporal expression pattern and functional analysis. Using morpholino oligo knockdown and mRNA overexpression, we demonstrated that zebrafish atg5 is required for normal morphogenesis of brain regionalization and body plan as well as for expression regulation of neural gene markers: gli1, huC, nkx2.2, pink1, β-synuclein, xb51 and zic1. We further demonstrated that ATG5 protein is involved in autophagy by LC3-II/LC3I ratio and rapamycin-induction experiments, and that ATG5 is capable of regulating expression of itself gene in the manner of a feedback inhibition loop. In addition, we found that expression of another autophagy-related gene, atg12, is maintained at a higher constant level like a housekeeping gene. This indicates that the formation of the ATG12-ATG5 conjugate may be dependent on ATG5 protein generation and its splicing, rather than on ATG12 protein in zebrafish. Importantly, in the present study, we provide a mechanistic insight into the regulation and functional roles of atg5 in development of zebrafish nervous system.

Autophagy is essential for cardiac morphogenesis during vertebrate development

“…All 4 transcripts were detected in 1-cell stage-embryos, indicating that they were maternally deposited, as well as at the sphere (3 hpf), germ ring (6 hpf) and 5-somite (12 hpf) stages and at 24 and 48 hpf. atg5 and becn1 transcripts appeared to decrease at the early gastrulation (germ ring) stage, similar to what was reported for atg5, 19 ambra1a, and ambra1b. 20 He and coworkers previously reported that several transcripts including ulk1b are not expressed before 23 hpf.…”

“…The result is aberrant development of the cardiac chambers and the atrioventricular canal, as well as increased embryonic lethality. While previous studies have indicated that specific autophagy factors are required for development of the Drosophila eye, 12,43 the zebrafish brain, 19,20 and for mouse development (reviewed in ref. 7), our results provide the first evidence that autophagy contributes to the cardiac morphogenic program during vertebrate development.…”

“…Hu and coworkers also identified atg5 transcripts at early developmental time points, including 0.2, 5, 10, 18, and 24 hpf. 19 To confirm the occurrence of autophagy in early embryonic development of zebrafish, 10-somite stage embryos were monitored for autophagosome structures by electron microscopy. We detected both early autophagosomes and autolysosomes in somite tissue (Fig.…”

“…Of note, in the zebrafish, Hu et al reported that knockdown of atg5 leads to abnormal morphogenesis of brain regionalization and body plan. 19 In contrast to mice lacking Atg3, Atg5, Atg7, Atg9, and Atg6l1, mice lacking 3 other genes involved in autophagy, Becn1, Ambra1, and Rb1cc1/Fip200, die during embryogenesis (reviewed in refs. 5 and 7).…”

At the Intersection of the Pathways for Exocytosis and Autophagy

“…Exocytosis and autophagy are essential for a number of common biological processes, including; the immune response (Govind, 2008;Minty et al, 1983;Murray et al, 1998;Ostenson et al, 2006), cell growth (Brennwald & Rossi, 2007;Orlando & Guo, 2009;Wei & Zheng, 2011;Zhang et al, 2005), cell proliferation and apoptosis (Kundu, 2011;Shin et al, 2011;Zeng et al, 2012), and multicellular organism development (Gutnick et al, 2011;Hu et al, 2011;Sato & Sato, 2011;Tra et al, 2011). Autophagy and exocytosis both involve membrane trafficking and fusion events and so similar groups of molecular machinery may be required for both processes: such as GTPase proteins, that facilitate membrane tethering and SNARE proteins which are involved in membrane fusion.…”