H3K27ac chromatin acetylation and gene expression analysis reveal sex- and situs-related differences in developing chicken gonads

Abstract

Background Birds exhibit a unique asymmetry in terms of gonad development. The female left gonad generates a functional ovary, whereas the right gonad regresses. In males, both left and right gonads would develop into testes. How is this left/right asymmetry established only in females but not in males remains unknown. The epigenetic regulation of gonadal developmental genes may contribute to this sex disparity. The modification of histone tails such as H3K27ac is tightly coupled to chromatin activation and gene expression. To explore whether H3K27ac marked chromatin activation is involved in the asymmetric development of avian gonads, we probed genome-wide H3K27ac occupancy in left and right gonads from both sexes and related chromatin activity profile to the expression of gonadal genes. Furthermore, we validated the effect of chromatin activity on asymmetric gonadal development by manipulating the chromatin histone acetylation levels. Methods The undifferentiated gonads from both sides of each sex were collected and subjected to RNA-Seq and H3K27ac ChIP-Seq experiments. Integrated analysis of gene expression and active chromatin regions were performed to identify the sex- and situs-specific regulation and expression of gonadal genes. The histone deacetylase inhibitor trichostatin A (TSA) was applied to the undifferentiated female right gonads to assess the effect of chromatin activation on gonadal gene expression and cell proliferation. Results Even before sex differentiation, the gonads already show divergent gene expression between different sexes and between left/right sides in females. The sex-specific H3K27ac chromatin distributions coincide with the higher expression of male/female specification genes in each sex. Unexpectedly, the H3K27ac marked chromatin activation show a dramatic difference between left and right gonads in both sexes, although the left/right asymmetric gonadal development was observed only in females but not in males. In females, the side-specific H3K27ac occupancy instructs the differential expression of developmental genes between the pair of gonads and contributes to the development of left but not right gonad. However, in males, the left/right discrepancy of H3K27ac chromatin distribution does not drive the side-biased gene expression or gonad development. The TSA-induced retention of chromatin acetylation causes up-regulation of ovarian developmental genes and increases cell proliferation in the female right gonad. Conclusions We revealed that left/right asymmetry in H3K27ac marked chromatin activation exists in both sexes, but this discrepancy gives rise to asymmetric gonadal development only in females. Other mechanisms overriding the chromatin activation would control the symmetric development of male gonads in chicken.

A PITX2–HTR1B pathway regulates the asymmetric development of female gonads in chickens

“…Pitx2 was shown to be downstream of Nodal activity, but it persists much longer throughout organogenesis than Nodal ( 39 ). For instance, Nodal is not expressed in chicken gonads, but Pitx2 is specifically expressed in the left gonads of both sexes ( 5 ). Thus, we set out to identify the signaling module conferring left-sided instructions to Pitx2 in the developing gonads, particularly in females.…”

“…From E6.5 onwards, the left gonads were growing and appearing bigger in size than the right one and the cortex of the left gonad is thicker than that in the right ( 1 ). Several reports studied this female-specific left/right asymmetrical development of gonads by mainly focusing on gene transcription ( 3 , 4 ) and chromatin activation ( 5 ). A couple of genes and related signaling pathways were distinctively expressed between left and right gonads.…”

Identification of oogonial stem cells in chicken ovary

“…It has long been accepted that in mammals, all oogonia enter meiosis during fetal life, with no remaining germ stem cells left in the ovary, in contrast to spermatogonia, which are present in the testis throughout life. 38 , 39 Nevertheless, the continuation of oogenesis into adulthood has been well documented in non‐mammalian species, including flies, 40 fishes, 14 amphibians 15 and reptiles. 16 This study provided the first evidence of the existence and functional relevance of OSCs in birds, which are considered to have undergone convergent evolution with mammals with respect to many physiological features.…”

“…It has long been accepted that in mammals, all oogonia enter meiosis during fetal life, with no remaining germ stem cells left in the ovary, in contrast to spermatogonia, which are present in the testis throughout life 38,39 . Nevertheless, the continuation of oogenesis into adulthood has been well documented in non‐mammalian species, including flies, 40 fishes, 14 amphibians 15 and reptiles 16 .…”

The Synchronized Progression from Mitosis to Meiosis in Female Primordial Germ Cells between Layers and Broilers

“…By E9.5, female and male gonads are morphologically and histologically disparate. In females, ovaries are known to develop asymmetrically, and only the left ovary develops a thick cortex where the germ cells are located, while the right one regresses [ 9 , 10 ].…”