AUTHOR=Menzies Brandon R. , Tarulli Gerard A. , Frankenberg Stephen R. , Pask Andrew J. 

TITLE=Therian origin of INSL3/RXFP2-driven testicular descent in mammals

JOURNAL=Frontiers in Cell and Developmental Biology

VOLUME=12

YEAR=2024

URL=https://www.frontiersin.org/journals/cell-and-developmental-biology/articles/10.3389/fcell.2024.1353598

DOI=10.3389/fcell.2024.1353598

ISSN=2296-634X

ABSTRACT=<p><bold>Introduction:</bold> During early development in most male mammals the testes move from a position near the kidneys through the abdomen to eventually reside in the scrotum. The transabdominal phase of this migration is driven by insulin-like peptide 3 (INSL3) which stimulates growth of the gubernaculum, a key ligament connecting the testes with the abdominal wall. While all marsupials, except the marsupial mole (<italic>Notoryctes typhlops</italic>), have a scrotum and fully descended testes, it is unclear if <italic>INSL3</italic> drives this process in marsupials especially given that marsupials have a different mechanism of scrotum determination and position relative to the phallus compared to eutherian mammals.</p><p><bold>Methods:</bold> To understand if INSL3 plays a role in marsupial testicular descent we have sequenced and curated the <italic>INSL3</italic> gene and its receptor (<italic>RXFP2</italic>) in a range of marsupials representing every order. Furthermore, we looked at single cell RNA-seq and qPCR analysis of INSL3 in the fat-tailed dunnart testis (<italic>Sminthopsis crassicaudata</italic>) to understand the location and timing of expression during development.</p><p><bold>Results:</bold> These data show a strong phylogenetic similarity between marsupial and eutherian orthologues, but not with monotreme INSL3s which were more similar to the ancestral RLN3 gene. We have also shown the genomic location of <italic>INSL3</italic>, and surrounding genes is conserved in a range of marsupials and eutherians. Single cell RNA-seq and qPCR data show that <italic>INSL3</italic> mRNA is expressed specifically in Leydig cells and expressed at higher levels during the testicular descent phase in developing marsupials.</p><p><bold>Discussion:</bold> Together, these data argue strongly for a therian origin of INSL3 mediated testicular descent in mammals and suggests that a coordinated movement of the testes to the abdominal wall may have preceded externalization in marsupials and therian mammals.</p>