Cellular Recycling Machine Linked to Male Fertility Hormone Control

Inside the testes, a population of supporting cells works tirelessly to shepherd developing sperm toward maturity. These cells, known as Sertoli cells, provide structural scaffolding, deliver nutrients, and secrete signaling molecules that germ cells need at every stage of their development. One of their most important products is androgen-binding protein, or ABP, a molecule that keeps testosterone concentrations high within the testis so that spermatogenesis can proceed. Although ABP has been studied for decades, a fundamental question has remained open: how do Sertoli cells get rid of ABP once it has done its job? A new study published in Reproductive Sciences by Ilkay Corumluoglu and Sevil Cayli of Ankara Yildirim Beyazit University offers the first evidence that a cellular recycling protein called p97/VCP is directly involved in the autophagic degradation of ABP, pointing to a previously unrecognized regulatory mechanism in male reproductive biology.

Autophagy is the cell’s housekeeping system, a process in which damaged proteins and organelles are engulfed in double-membraned vesicles called autophagosomes and delivered to lysosomes for destruction. Two molecular markers define this pathway: LC3, a protein that decorates the autophagosomal membrane, and p62, an adaptor that ferries ubiquitinated cargo to the growing autophagosome. The AAA-ATPase p97/VCP, meanwhile, is a hexameric molecular motor best known for extracting ubiquitinated proteins from cellular structures and handing them off to the proteasome or the autophagy machinery. Previous work by the same group had shown that blocking p97/VCP function in mouse Sertoli cells causes defective autophagosome maturation, cell cycle arrest, and apoptosis, making the protein a plausible candidate for regulating hormone-related protein turnover in these cells.

To test this idea, the researchers used the Sertoli 15P-1 cell line, a well-established mouse model derived through transmeiotic differentiation of male germ cells in culture. They exposed the cells to three pharmacological agents chosen for their opposing effects on autophagic flux. Testosterone served as the physiological ligand relevant to ABP regulation. Chloroquine, a lysosomotropic agent, blocks the fusion of autophagosomes with lysosomes, causing LC3 and p62 to accumulate because cargo can no longer be degraded. Rapamycin, by contrast, inhibits the mTOR signaling pathway and stimulates autophagy, accelerating the clearance of cellular components. Comparing the effects of these treatments allowed the team to infer whether ABP and p97/VCP travel together through the autophagic pipeline.

The experimental design combined three complementary techniques. Western blotting quantified protein expression levels across treatment groups, providing a biochemical readout of pathway activity. Single and double immunofluorescence staining revealed where p97/VCP, ABP, LC3, and p62 reside within the cells and, crucially, whether their locations overlap, a phenomenon called colocalization that suggests functional interaction. Finally, small interfering RNA (siRNA) transfection was used to specifically silence the gene encoding p97/VCP, creating a loss-of-function condition against which the drug treatments could be compared. This triangulation of methods strengthens the interpretation, because each approach has distinct limitations that the others can compensate for.

The Western blot results painted a picture consistent with autophagic involvement. p62 expression was highest in the group treated with both chloroquine and testosterone compared with the control siRNA condition, although the difference did not reach statistical significance. LC3 expression peaked in the chloroquine-treated group, again without reaching significance, which is the expected pattern when autophagic flux is blocked at the lysosomal step: autophagosomes accumulate because they cannot be cleared. As anticipated, the lowest expression of p97/VCP was detected in the VCPsi group, confirming that the siRNA transfection successfully knocked down the target protein and validating the loss-of-function arm of the study.

The immunofluorescence analyses delivered the study’s most striking findings. Significant differences in protein colocalization patterns emerged across all treatment groups, with one notable exception: p62 in rapamycin-treated samples showed no significant change. This pattern suggests that the physical association between p97/VCP and ABP shifts depending on the state of the autophagic machinery. When chloroquine jams the lysosomal bottleneck, proteins destined for degradation pile up and their colocalization with p97/VCP changes; when rapamycin revs up autophagy, the dynamics of interaction shift again. The authors report that these data provide the first evidence in the literature of a direct relationship between p97/VCP and ABP within Sertoli cells, a finding that reframes how scientists think about hormonal regulation in the testis.

Why does this matter for fertility? ABP’s job is to bind testosterone and maintain the extraordinarily high intratesticular concentrations that developing germ cells require. Disrupting ABP levels has measurable consequences: transgenic mice overexpressing ABP show meiotic arrest and germ cell apoptosis, and altered ABP expression has been linked to impaired sperm function in multiple models. If p97/VCP governs how quickly ABP is degraded, then the recycling motor effectively acts as a dial controlling how much ABP, and by extension how much bioavailable testosterone, is present in the seminiferous tubule environment at any given time. The data suggest a novel regulatory mechanism by which p97/VCP may influence spermatogenesis and male fertility through ABP degradation, offering a potential explanation for some forms of testicular dysfunction that currently lack a molecular diagnosis.

The implications extend well beyond the testis. ABP is not exclusive to Sertoli cells; it is also expressed in the brain, the liver, and the placenta, tissues where steroid hormone handling is equally critical. The authors propose that their work lays a foundational framework for future investigations in these ABP-expressing tissues, potentially linking p97/VCP expression to endocrine and pathophysiological disorders far removed from reproduction. Notably, p97/VCP has already been implicated in preeclamptic placenta, where altered expression coincides with impaired autophagy, and in testicular ischemia-reperfusion injury. Mutations in the VCP gene also cause multisystem proteinopathy, a devastating condition combining muscle, bone, and neurodegenerative disease, underscoring how central this protein is to cellular protein quality control across organ systems.

Technically, the study sits at the intersection of two major protein degradation systems. p97/VCP was originally identified as an ATPase co-purified with the inhibitor of NF-kappaB and the 26S proteasome, and it remains a linchpin of ubiquitin-proteasome-mediated degradation. But accumulating evidence shows that p97/VCP also cooperates with autophagy: in yeast, the orthologous Cdc48 protein regulates autophagosome biogenesis in concert with the ubiquitin-like protein Atg8, and in mammalian cells p97 inhibitors impair both ubiquitin-dependent and autophagic protein clearance. The p62 adaptor itself bridges the two systems, linking the autophagy pathway to the ubiquitin-proteasome system during the degradation of ubiquitinated proteins. The new findings place ABP squarely within this dual-disposal network in Sertoli cells.

As with any cell-line study, caveats apply. The statistical comparisons for p62 and LC3 did not reach significance, so the biochemical trends should be treated as suggestive rather than definitive, and the colocalization data, while significant, demonstrate proximity rather than direct biochemical binding. Follow-up work in primary Sertoli cells, intact seminiferous tubules, and animal models will be needed to confirm that the p97/VCP-ABP axis operates in vivo and to determine whether manipulating it can rescue fertility in disease settings. Nevertheless, by connecting a master regulator of protein homeostasis to a keystone hormone-binding protein, the study opens a concrete new avenue for research into male infertility, and it hints that therapies targeting autophagy, a strategy already being explored in fields from neurodegeneration to cancer, may one day find applications in reproductive medicine as well.

Subject of Research: Autophagic degradation of androgen-binding protein by p97/VCP in Sertoli cells and its implications for male fertility

Article Title: The Autophagic Interaction Between p97/VCP and ABP Protein Expressions in Sertoli Cell Line: the Effects of Chloroquine and Rapamycin

Article References: Corumluoglu, I., & Cayli, S. (2026). The Autophagic Interaction Between p97/VCP and ABP Protein Expressions in Sertoli Cell Line: the Effects of Chloroquine and Rapamycin. Reproductive Sciences. https://doi.org/10.1007/s43032-026-02231-3

Image Credits: AI Generated

DOI: 10.1007/s43032-026-02231-3

Keywords: p97/VCP, androgen-binding protein, autophagy, Sertoli cells, spermatogenesis, testosterone, chloroquine, rapamycin, male infertility, siRNA, LC3, p62