Abstract
Purpose :
Hypoxia-inducible factor 1 (HIF1α) is a critical transcription factor known to govern vascularization. The lens contains an oxygen gradient that parallels the surface to core differentiation of lens epithelial cells into fiber cells. The presence of this oxygen gradient suggests that HIF1α could mediate gene expression events required for lens cell differentiation and fiber cell function. Consistently, we have recently identified HIF1α to be an essential regulator of BNIP3L that we have shown initiates the hypoxia-dependent elimination of non-nuclear organelles during fiber cell maturation. Here, we employed a multiomics approach to map the genomic complement of HIF1α DNA binding sites and target genes in the lens and identify HIF1α as a novel master regulator of lens gene expression.
Methods :
HIF1α DNA binding was induced in embryonic primary chick lens cells through exposure to dimethyloxallyl glycine (DMOG). HIF1α-DNA binding complexes were identified by CUT&RUN (Cleavage Under Targets and Release Under Nuclease). Expression levels of activated or repressed HIF1α-target genes were identified by parallel RNA sequencing analysis. Sites were mapped to putative promoter or enhancer sequences and analyzed for chromatin configuration by analysis of ATAC sequencing data.
Results :
CUT&RUN analysis revealed 8,375 HIF1α-DNA binding complexes across the chick lens genome. 1,190 HIF1α-DNA binding complexes were significantly clustered within open chromatin regions (p < 1e-55) identified by ATAC sequencing. Formation of the identified HIF1α-DNA complexes parallels the direct activation or repression of 526 genes, 202 of which contained HIF1α binding sites within 100kbp of the transcription start site. The identified genes were associated with multiple key cellular processes including glycolysis, cell cycle control, chromatin remodeling, Notch and Wnt signaling, lens cell differentiation, development, and cataract formation.
Conclusions :
The data establish a functional genomic map of novel HIF1α-regulated lens genes. They further support the hypothesis that hypoxia is critical for lens fiber cell differentiation, structure and function and they identify novel HIF1α-dependent pathways and genetic components important for lens differentiation, homeostasis and cataract formation.
This is a 2021 ARVO Annual Meeting abstract.