Functions of HIF Gene Regulatory Network in Zebrafish Neural Development

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基本資料

Project title

Code/計畫編號

NSC102-2313-B019-010-MY3

Translated Name/計畫中文名

HIF基因調控網路在斑馬魚神經發育過程中的角色

Project Coordinator/計畫主持人

Chin-Hwa Hu

Funding Organization/主管機關

National Science and Technology Council

Co-Investigator(s)/共同執行人

白敦文
鄒文雄

Department/Unit

Department of Bioscience and Biotechnology

Website

https://www.grb.gov.tw/search/planDetail?id=8104717

Year

2014

Start date/計畫起

01-08-2014

Expected Completion/計畫迄

01-07-2015

Co-Investigator(s)

Wen-Shyong Tzou

Bugetid/研究經費

1420千元

ResearchField/研究領域

生物技術(理)
生物科學

氧氣是維持生命不可或缺的重要分子，當遇到低氧的緊迫環境時，細胞會藉由低氧訊息傳遞系統的低氧誘發蛋白HIF（包括HIF1α~-3α及HIFβ），啟動生物適應相關基因，改變細胞內的生理代謝途徑，藉以提供細胞所需的能量或增加氧氣的運輸能力。過去長期以來，大部份的研究一直將細胞低氧訊息系統視為生物調適低氧逆境的一環，但是有關於這套細胞訊息系統在正常供氧環境下的生理功能則較少被探討。我們最近的研究發現雖然在正常（常氧）環境下細胞內的HIF1α與HIF2α表現量低，但是對斑馬魚胚胎的發育卻有許多重要的功能，例如保護神經細胞促進分化、統整造血系統的細胞生長與分化、促進視網膜與肝臟發育等息息相關。將hif2α默化會降低神經先趨細胞內的抗凋亡基因survivin/birc5a, -5b的表現，使得細胞大量凋亡，同時會使得未分化的神經先趨細胞無法脫離細胞週期進入神經分化的過程。因此在胚胎的中樞神經細胞發育過程中，HIF2α巧妙地藉由控制細胞凋亡抑制蛋白基因的表現，阻止神經先驅細胞凋亡且同時驅動神經細胞的分化工作。除此之外，hif2α默化亦會抑制神經細胞的內多種neurod基因的表現，並造成視網膜神經細胞的分化受到抑制。外源性nrd cRNA或是birc5a/-5b cRNA均可以成功恢復hif2α默化的斑馬魚胚胎內視網膜感光細胞的分化，顯示HIF2α、Survivin/ Birc5a, -5b、與NeuroD三者之間在視網膜的發育上有著密切的相互關聯性，但是其機制尚不清楚。另外最近的研究發現兩個神經細胞的攜氧球蛋白基因Neuroglobin (Ngb)與Cytoglobin (Cygb)，分別會受到低氧緊迫訊息系統的誘發而大量表現，可能與供應細胞氧氣、清除自由基、以及保護細胞避免凋亡有關。在本計畫中我們想深入探討在胚胎中樞神經與視網膜神經發育過程中，HIF2α、Birc5a/-5b、NeuroD家族、以及受低氧訊息控制的Ngb/Cygb彼此之間究竟存在什麼樣的關係？各自扮演什麼樣的角色？我們將分別從個別基因組功能以及整體基因調控網路的角度探討這些基因群對神經發育的功能。我們將利用反向遺傳與基因轉錄體分析的方法，透過基因默化與救援、定性與定量基因表現活性分析、高通量基因轉錄體定序、基因本體論分析以及染色體免疫共沉澱等實驗與分析，探討在神經發育過程中低氧誘發蛋白HIF2α與其下游Birc5a/-5b、NeuroD家族以及Ngb/Cygb之間相互的關聯和生物功能的互補性，並建立胚胎內hif2α、nrd家族、birc5a/-5b、ngb/cygb的相關基因調控網路。另將藉由比對常/低氧環境下控制組及hif2α默化胚胎內基因轉錄體的差異，探討HIF2α在正常與低氧緊迫環境下所扮演的生物功能。 Oxygen is the most important molecule in our life, which can provide pivotal biological energy through an aerobic respiration system. When the organism is under hypoxia stress, it will make some physiological changes through a hypoxia-inducible factors (HIFs)-mediated signaling system. The HIFs are heterodimeric transcription factors consisting with an oxygen-regulated α subunit and a constitutively expressed β subunit (HIF1β/ARNT). When cells are subjected to hypoxia, the HIFα (HIF1α and HIF2α) factors are stabilized and in turn associate with HIF1β/ARNT and activate target genes. Frequently, the cellular hypoxia signaling pathway is regarded as a stress-response system to protect the organism itself when it is subjected to hypoxia. Our recent studies, however, revealed that the cellular hypoxia-signaling system plays critical functions during zebrafish development, including neural survival and differentiation, erythropoiesis, liver outgrowth, and retinal development. It suggests that even with low level of HIFα factors existed, the hypoxia-signaling system still plays critical biological functions in normoxia environment. HIF2α depletion caused massive cell death and abrogated neural cell differentiation due to aberrant expression of the survivin homologues (birc5a and birc5b). It appears that Birc5a/-5b act downstream of HIF2α to protect neural progenitor cells and promote neural differentiation. In addition, we found that HIF2α depletion also repressed expression of neurod family and inhibited retinal differentiation. Ectopic nrd or birc5a/-5b cRNAs in hif2α morphant could rescue photoreceptor differentiation successfully. It suggests a close interaction network between hif2α, nrd, and birc5a/-5b genes during development. Recent studies demonstrated that the neural globin genes, neuroglobin/ngb and cytoglobin/cygb, can be induced by hypoxia treatment. Both Ngb and Cygb involve in neural protection, ROS scavenging, and oxygen homeostasis. With the core techniques of reverse genetics, genomics, and bioinformatics (including gene silencing and rescuing, quantitative and qualitative gene expression analysis, high throughput RNA sequencing, gene ontology, chromatin immunoprecipitation techniques), here we would like to investigate the correlations between hif2α, nrd, birc5a/-5b, and ngb/cygb during CNS and retinal development. Accordingly, we would like to establish a hif2α-related functional and gene regulation network during neural development. Furthermore, we would like to investigate the functions of HIF2α-mediated hypoxia-signaling system in normoxia- and hypoxia-conditions, with respect to the genetic regulation network,

Keyword(s)

斑馬魚胚胎
低氧誘發蛋白HIF2α
抗凋亡蛋白Birc5a/-5b
神經攜氧蛋白Ngb/Cygb
神經細胞基因調控蛋白NeuroD
神經發育
基因轉錄體
基因默化
HIF2α
Birc5a/-5b
NeuroD
neuroglobin
cytoglobin
neural development
transcriptome
gene silencing
zebrafish

DSpace CRIS

Functions of HIF Gene Regulatory Network in Zebrafish Neural Development

基本資料

Description