Center for Cancer Research, Massachusetts General Hospital, Charlestown, Massachusetts, USA.
Department of Medicine, Harvard Medical School, Boston, Massachusetts, USA.
Endocrinology. 2021 Jan 1;162(1). doi: 10.1210/endocr/bqaa208.
Whereas in ovo exposure of genetically male (ZZ) chicken embryos to exogenous estrogens temporarily feminizes gonads at the time of hatching, the morphologically ovarian ZZ-gonads (FemZZs for feminized ZZ gonads) are masculinized back to testes within 1 year. To identify the feminization-resistant "memory" of genetic male sex, FemZZs showing varying degrees of feminization were subjected to transcriptomic, DNA methylome, and immunofluorescence analyses. Protein-coding genes were classified based on their relative mRNA expression across normal ZZ-testes, genetically female (ZW) ovaries, and FemZZs. We identified a group of 25 genes that were strongly expressed in both ZZ-testes and FemZZs but dramatically suppressed in ZW-ovaries. Interestingly, 84% (21/25) of these feminization-resistant testicular marker genes, including the DMRT1 master masculinizing gene, were located in chromosome Z. Expression of representative marker genes of germline cells (eg, DAZL or DDX4/VASA) was stronger in FemZZs than normal ZZ-testes or ZW-ovaries. We also identified 231 repetitive sequences (RSs) that were strongly expressed in both ZZ-testes and FemZZs, but these RSs were not enriched in chromosome Z. Although 94% (165/176) of RSs exclusively expressed in ZW-ovaries were located in chromosome W, no feminization-inducible RS was detected in FemZZs. DNA methylome analysis distinguished FemZZs from normal ZZ- and ZW-gonads. Immunofluorescence analysis of FemZZ gonads revealed expression of DMRT1 protein in medullary SOX9+ somatic cells and apparent germline cell populations in both medulla and cortex. Taken together, our study provides evidence that both somatic and germline cell populations in morphologically feminized FemZZs maintain significant transcriptomic and epigenetic memories of genetic sex.
虽然在鸡胚孵化时，向雄性（ZZ）胚胎中暴露外源性雌激素会暂时使生殖腺女性化，但形态上的卵巢 ZZ 生殖腺（FemZZ，女性化的 ZZ 生殖腺）在 1 年内会被雄性化回睾丸。为了确定遗传雄性的抗雌性化“记忆”，我们对表现出不同程度雌性化的 FemZZ 进行了转录组、DNA 甲基化组和免疫荧光分析。根据它们在正常 ZZ 睾丸、遗传上的雌性（ZW）卵巢和 FemZZ 中的相对 mRNA 表达，将蛋白质编码基因进行了分类。我们鉴定了一组 25 个基因，它们在 ZZ 睾丸和 FemZZ 中都强烈表达，但在 ZW 卵巢中被显著抑制。有趣的是，这些抗雌性化的睾丸标记基因中有 84%（21/25），包括 DMRT1 主要的雄性化基因，位于染色体 Z 上。生殖细胞的代表性标记基因（如 DAZL 或 DDX4/VASA）在 FemZZ 中的表达比正常 ZZ 睾丸或 ZW 卵巢更强。我们还鉴定了 231 个重复序列（RS），它们在 ZZ 睾丸和 FemZZ 中都强烈表达，但这些 RS 不在染色体 Z 上富集。尽管 94%（165/176）仅在 ZW 卵巢中表达的 RS 位于染色体 W 上，但在 FemZZ 中未检测到雌性化诱导的 RS。DNA 甲基化组分析将 FemZZ 与正常的 ZZ 和 ZW 生殖腺区分开来。FemZZ 生殖腺的免疫荧光分析显示，DMRT1 蛋白在髓质 SOX9+体细胞和皮质和髓质中的明显生殖细胞群体中表达。总之，我们的研究提供了证据，表明形态上女性化的 FemZZ 中的体细胞和生殖细胞群体都保持着遗传性别显著的转录组和表观遗传记忆。
