Single-Cell RNA Sequencing Analysis of Chicken Anterior Pituitary: A Bird’s-Eye View on Vertebrate Pituitary

Abstract

It is well-established that anterior pituitary contains multiple endocrine cell populations, and each of them can secrete one/two hormone(s) to regulate vital physiological processes of vertebrates. However, the gene expression profiles of each pituitary cell population remains poorly characterized in most vertebrate groups. Here we analyzed the transcriptome of each cell population in adult chicken anterior pituitaries using single-cell RNA sequencing technology. The results showed that: (1) four out of five known endocrine cell clusters have been identified and designated as the lactotrophs, thyrotrophs, corticotrophs, and gonadotrophs, respectively. Somatotrophs were not analyzed in the current study. Each cell cluster can express at least one known endocrine hormone, and novel marker genes (e.g., CD24 and HSPB1 in lactotrophs, NPBWR2 and NDRG1 in corticotrophs; DIO2 and SOUL in thyrotrophs, C5H11ORF96 and HPGDS in gonadotrophs) are identified. Interestingly, gonadotrophs were shown to abundantly express five peptide hormones: FSH, LH, GRP, CART and RLN3; (2) four non-endocrine/secretory cell types, including endothelial cells (expressing IGFBP7 and CFD) and folliculo-stellate cells (FS-cells, expressing S100A6 and S100A10), were identified in chicken anterior pituitaries. Among them, FS-cells can express many growth factors, peptides (e.g., WNT5A, HBEGF, Activins, VEGFC, NPY, and BMP4), and progenitor/stem cell-associated genes (e.g., Notch signaling components, CDH1), implying that the FS-cell cluster may act as a paracrine/autocrine signaling center and enrich pituitary progenitor/stem cells; (3) sexually dimorphic expression of many genes were identified in most cell clusters, including gonadotrophs and lactotrophs. Taken together, our data provides a bird’s-eye view on the diverse aspects of anterior pituitaries, including cell composition, heterogeneity, cell-to-cell communication, and gene expression profiles, which facilitates our comprehensive understanding of vertebrate pituitary biology.

A Gene Expression Atlas of Lohmann White Chickens

“…RLN3 is highly expressed in female pituitary gonadotrophs but not in male pituitary gonadotrophs. Similarly, GRP , HPGDS , and CPLX1 have relatively higher mRNA levels in female than that in male gonadotrophs, which was confirmed by using quantitative real-time PCR (qPCR) to indicate the sexually dimorphic expression patterns of GRP , RLN3 , and HPGDS at the whole anterior pituitary level (Zhang et al, 2021). In addition, some novel differentially expressed genes were revealed in this study.…”

“…In the chicken pituitary, some genes with a high TPM value (>200) are expressed in a sex-dependent manner, including LOC121107016 , LOC121107017 , LOC112533599 , CPLX1 , RLN3 , GRP , HPGDS , HEXB , SPARCL1 in female, and HSPA5 , PLK2 , CSRP2 , CCDC80 , LOC112532140 , MEM201 in male (Figure 6E). Interestingly, in our previous scRNA-seq study, we found that in gonadotrophs clusters of the chicken anterior pituitary, some genes ( CPLX1 , RLN3 , GRP and HPGDS ) are also expressed in a sex-dependent manner (Zhang et al, 2021). RLN3 is highly expressed in female pituitary gonadotrophs but not in male pituitary gonadotrophs.…”

Characterization of CRH-Binding Protein (CRHBP) in Chickens: Molecular Cloning, Tissue Distribution and Investigation of Its Role as a Negative Feedback Regulator within the Hypothalamus–Pituitary–Adrenal Axis

“…Especially, the mouse CRHBP is detected to be highly expressed in the pituitary gland with a sexually dimorphic pattern, with females exhibiting much higher expression levels [ 30 , 50 ]. Single-cell sequencing from our research group further reveals that chicken CRHBP is abundantly expressed in pituitary folliculostellate cells (FS cells) [ 51 ], being different from the detection in female mice that CRHBP is expressed in pituitary cells, including corticotropes, lactotropes and gonadotropes [ 30 ]. The FS cells in the chicken pituitary are hypothesized to function as a paracrine/autocrine signaling hub, influencing neighboring cells [ 51 ].…”

Characterization of the chicken melanocortin 5 receptor and its potential role in regulating hepatic glucolipid metabolism

“…cDNA samples were subjected to quantitative real-time PCR assay of chicken MC5R mRNA levels in different chicken tissues, as described in our previous study ( Fang et al, 2021 ; Zhang et al, 2021 ). Quantitative real-time PCR (qPCR) was performed on the CFX96 Real-time PCR Detection System (Bio-Rad, Hercules, CA).…”