About

Prabhakara Reddi is a Professor in the Department of Comparative Biosciences at the University of Illinois College of Veterinary Medicine. His research focuses on understanding the mechanisms regulating mammalian spermatogenesis, with the long-term goal of developing novel therapies for male infertility, which is a significant health concern affecting approximately 15% of couples of reproductive age. His work investigates the molecular regulation of spermatogenesis, including transcriptional regulation and the role of specific proteins such as TDP-43 in male germ cell differentiation and fertility. Professor Reddi has contributed to elucidating how RNA Polymerase II pausing maintains precise gene expression patterns within the seminiferous epithelium and how insulator-mediated repression ensures testis-specific genes remain silent in somatic tissues. His research also explores the function of TDP-43 as a transcriptional repressor in male germ cells, its recruitment to promoters, and its importance for spermatogenesis and male fertility. His findings have implications for understanding infertility and the molecular basis of germ cell development.

Research topics

• Biology
• Cell biology
• Neuroscience
• Medicine
• Computational biology
• Pathology
• Endocrinology
• Ecology
• Genetics
• Evolutionary biology

Selected publications

• Transillumination-Assisted Microdissection for Precise Staging of Seminiferous Tubules in Mice

  Methods in molecular biology · 2025-01-01

  articleSenior author
  PublisherDOI

• RNA polymerase II pausing is essential during spermatogenesis for appropriate gene expression and completion of meiosis

  Nature Communications · 2024 · 24 citations

  Senior authorCorresponding
  • Genetics
  • Biology
  • Cell biology

  Male germ cell development requires precise regulation of gene activity in a cell-type and stage-specific manner, with perturbations in gene expression during spermatogenesis associated with infertility. Here, we use steady-state, nascent and single-cell RNA sequencing strategies to comprehensively characterize gene expression across male germ cell populations, to dissect the mechanisms of gene control and provide new insights towards therapy. We discover a requirement for pausing of RNA Polymerase II (Pol II) at the earliest stages of sperm differentiation to establish the landscape of gene activity across development. Accordingly, genetic knockout of the Pol II pause-inducing factor NELF in immature germ cells blocks differentiation to spermatids. Further, we uncover unanticipated roles for Pol II pausing in the regulation of meiosis during spermatogenesis, with the presence of paused Pol II associated with double-strand break (DSB) formation, and disruption of meiotic gene expression and DSB repair in germ cells lacking NELF.

  PublisherOA PDFDOI

• Transcriptional regulation of spatiotemporal gene expression within the seminiferous epithelium: Mouse <i>Acrv1</i> gene as a model

  Andrology · 2023-02-16 · 6 citations

  reviewOpen access1st authorCorresponding

  Precise spatiotemporal expression of cohorts of differentiation markers unique to spermatogonia, spermatocytes, and round spermatids punctuates spermatogenesis and ensures its completion. For example, genes coding for the synaptonemal complex or the acrosome or flagellum are expressed sequentially in a developmental stage- and germ cell-specific manner. But the transcriptional mechanisms governing the spatiotemporal order of gene expression within the seminiferous epithelium are poorly understood. Using the round spermatid-specific Acrv1 gene, which codes for the acrosomal protein SP-10 as a model, we learned that (1) the proximal promoter itself contains all the necessary cis-regulatory sequences, (2) an insulator prevents somatic cell expression of the testis-specific gene, (3) RNA II polymerase is loaded on the Acrv1 promoter but paused in spermatocytes, thus ensuring precise transcriptional elongation in round spermatids, and that (4) a transcriptional repressor binding protein of 43 kilodaltons (TDP-43) plays a role in maintaining the paused state in spermatocytes. Although the Acrv1 enhancer element has been narrowed down to 50 bp and its binding to a 47 kDa testis-abundant nuclear protein shown, the identity of the putative transcription factor responsible for activation of round spermatid-specific transcription remains elusive. Human male infertility is idiopathic with limited treatment options. Understanding transcriptional regulation of spermatogenesis has the potential to lead to future therapies for male infertility.

  PublisherOA PDFDOI

• RNA polymerase II pausing is essential during spermatogenesis for appropriate gene expression and completion of meiosis

  bioRxiv (Cold Spring Harbor Laboratory) · 2023-05-09 · 6 citations

  preprintOpen accessSenior authorCorresponding

  Male germ cell development requires precise regulation of gene activity in a cell-type and stage-specific manner, with perturbations in gene expression during spermatogenesis associated with infertility. Here, we use steady-state, nascent and single-cell RNA sequencing strategies to comprehensively characterize gene expression across male germ cell populations, to dissect the mechanisms of gene control and provide new insights towards therapy. We discover a requirement for pausing of RNA Polymerase II (Pol II) at the earliest stages of sperm differentiation to establish the landscape of gene activity across development. Accordingly, genetic knockout of the Pol II pause-inducing factor NELF in immature germ cells blocks differentiation to mature spermatids. Further, we uncover unanticipated roles for Pol II pausing in the regulation of meiosis during spermatogenesis, with the presence of paused Pol II associated with double strand break formation by SPO11, and disruption of SPO11 expression in germ cells lacking NELF.

  PublisherOA PDFDOI

• Sertoli cells require TDP-43 to support spermatogenesis

  Biology of Reproduction · 2022-08-20 · 6 citations

  articleOpen accessSenior authorCorresponding

  TAR DNA binding protein of 43 kD (TDP-43) is an evolutionarily conserved, ubiquitously expressed transcription factor and RNA-binding protein with major human health relevance. TDP-43 is present in Sertoli and germ cells of the testis and is aberrantly expressed in the sperm of infertile men. Sertoli cells play a key role in spermatogenesis by offering physical and nutritional support to male germ cells. The current study investigated the requirement of TDP-43 in Sertoli cells. Conditional knockout (cKO) of TDP-43 in mouse Sertoli cells caused failure of spermatogenesis and male subfertility. The cKO mice showed decreased testis weight, and low sperm count. Testis showed loss of germ cell layers, presence of vacuoles, and sloughing of round spermatids, suggesting loss of contact with Sertoli cells. Using a biotin tracer, we found that the blood-testis barrier (BTB) was disrupted as early as postnatal day 24 and worsened in adult cKO mice. We noted aberrant expression of the junction proteins connexin-43 (gap junction) and N-cadherin (ectoplasmic specialization). Oil Red O staining showed a decrease in lipid droplets (phagocytic function) in tubule cross-sections, Sertoli cells cytoplasm, and in the lumen of seminiferous tubules of cKO mice. Finally, qRT-PCR showed upregulation of genes involved in the formation and/or maintenance of Sertoli cell junctions as well as in the phagocytic pathway. Sertoli cells require TDP-43 for germ cell attachment, formation and maintenance of BTB, and phagocytic function, thus indicating an essential role for TDP-43 in the maintenance of spermatogenesis.

  PublisherOA PDFDOI

• Loss of TDP-43 in male germ cells causes meiotic failure and impairs fertility in mice

  Journal of Biological Chemistry · 2021-09-29 · 17 citations

  articleOpen accessSenior authorCorresponding

  Meiotic arrest is a common cause of human male infertility, but the causes of this arrest are poorly understood. Transactive response DNA-binding protein of 43 kDa (TDP-43) is highly expressed in spermatocytes in the preleptotene and pachytene stages of meiosis. TDP-43 is linked to several human neurodegenerative disorders wherein its nuclear clearance accompanied by cytoplasmic aggregates underlies neurodegeneration. Exploring the functional requirement for TDP-43 for spermatogenesis for the first time, we show here that conditional KO (cKO) of the Tardbp gene (encoding TDP-43) in male germ cells of mice leads to reduced testis size, depletion of germ cells, vacuole formation within the seminiferous epithelium, and reduced sperm production. Fertility trials also indicated severe subfertility. Spermatocytes of cKO mice showed failure to complete prophase I of meiosis with arrest at the midpachytene stage. Staining of synaptonemal complex protein 3 and γH2AX, markers of the meiotic synaptonemal complex and DNA damage, respectively, and super illumination microscopy revealed nonhomologous pairing and synapsis defects. Quantitative RT-PCR showed reduction in the expression of genes critical for prophase I of meiosis, including Spo11 (initiator of meiotic double-stranded breaks), Rec8 (meiotic recombination protein), and Rad21L (RAD21-like, cohesin complex component), as well as those involved in the retinoic acid pathway critical for entry into meiosis. RNA-Seq showed 1036 upregulated and 1638 downregulated genes (false discovery rate <0.05) in the Tardbp cKO testis, impacting meiosis pathways. Our work reveals a crucial role for TDP-43 in male meiosis and suggests that some forms of meiotic arrest seen in infertile men may result from the loss of function of TDP-43.

  PublisherDOI

• Spinal muscular atrophy: Broad disease spectrum and sex-specific phenotypes

  Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease · 2021 · 47 citations

  • Biology
  • Pathology
  • Medicine
  PublisherOA PDFDOI

• Mouse Sertoli cells isolation by lineage tracing and sorting

  Molecular Reproduction and Development · 2020-07-31 · 6 citations

  articleSenior authorCorresponding

  Sertoli cells play a key role in spermatogenesis by supporting the germ cells throughout differentiation. The isolation of Sertoli cells is essential to study their functions. However, the close contact of Sertoli cells with other testicular cell types and the high proliferation of contaminating cells are obstacles to obtain pure primary cultures. Current rodent Sertoli cell isolation protocols result in enriched, rather than pure Sertoli cells. Therefore, novel approaches are necessary to improve the purity of Sertoli cell primary cultures. The goal of this study is to obtain pure mouse Sertoli cells using lineage tracing and fluorescence-activated cell sorting (FACS). We bred the Amh-Cre mouse line with tdTomato line to generate mice constitutively expressing red fluorescence specifically in Sertoli cells. Primary cultures of Sertoli cells isolated from prepubertal mice showed that 79% of cells expressed tdTomato, as evaluated by fluorescence microscopy and flow cytometry; however, nearly all adherent cells were positive for vimentin. Most of the tomato-negative cells expressed α-smooth muscle actin (α-SMA), a peritubular myoid cell marker, but double-negative populations were also present. These findings suggest that vimentin lacks Sertoli cell-specificity and that α-SMA is not adequate to identify all of the contaminating cells. Upon FACS sorting; however, virtually 100% of the cells were tdTomato positive, expressed vimentin, but not α-SMA. Prepubertal mice yielded a higher number of Sertoli cells compared to adults, but both could be adequately sorted. In conclusion, our study shows that lineage tracing and sorting is an efficient strategy for acquiring pure populations of murine Sertoli cells.

  PublisherDOI

• Acrosomal marker SP-10 (gene name Acrv1) for staging of the cycle of seminiferous epithelium in the stallion

  Theriogenology · 2020-07-06 · 12 citations

  articleOpen accessSenior authorCorresponding
  PublisherOA PDFDOI

• Characterization of rodent Sertoli cell primary cultures

  Molecular Reproduction and Development · 2020 · 33 citations

  Senior authorCorresponding
  • Biology
  • Evolutionary biology
  • Cell biology

  Sertoli cells play a vital role in spermatogenesis by offering physical and nutritional support to the differentiating male germ cells. They form the blood-testis barrier and secrete growth factors essential for germ cell differentiation. Sertoli cell primary cultures are critical for understanding the regulation of spermatogenesis; however, obtaining pure cultures has been a challenge. Rodent Sertoli cell isolation protocols do not rule out contamination by the interstitial or connective tissue cells. Sertoli cell-specific markers could be helpful, but there is no consensus. Vimentin, the most commonly used marker, is not specific for Sertoli cells since its expression has been reported in peritubular myoid cells, mesenchymal stem cells, fibroblasts, macrophages, and endothelial cells, which contaminate Sertoli cell preparations. Markers based on transcription and growth factors also have limitations. Thus, the impediment to obtaining pure Sertoli cell cultures pertains to both the method of isolation and marker usage. The aim of this review is to discuss improvements to current methods of rodent Sertoli cell primary cultures, assess the properties of prepubertal versus mature Sertoli cell cultures, and propose steps to improve cellular characterization. Potential benefits of using contemporary approaches, including lineage tracing, specific cell ablation, and RNA-seq for obtaining Sertoli-specific transcript markers are discussed. Evaluating the specificity and applicability of these markers at the protein level to characterize Sertoli cells in culture would be critical. This review is expected to positively impact future work using primary cultures of rodent Sertoli cells.

  PublisherOA PDFDOI

Recent grants

• NIH Grant R01HD036239

  NIH · $1.5M · 2010

• NIH Grant R29HD036239

  NIH · $520k · 2004

• RNA Pol II Pausing is Critical for Spermatogenesis and Male Fertility

  NIH · $1.7M · 2018–2025

Frequent coauthors

• John C. Herr

  University of Virginia

  15 shared

• Charles J. Flickinger

  University of Virginia

  8 shared

• Pramod Khandekar

  6 shared

• Helena D. Zomer

  University of Florida

  6 shared

• Michael J. Wolkowicz

  University of Virginia

  5 shared

• Craig Urekar

  University of Virginia

  5 shared

• Mayuresh M. Abhyankar

  University of Virginia

  4 shared

• Amy N. Shore

  Biomedical Research Institute

  4 shared

Education

• Research Instructor, Cell Biology

  University of Virginia School of Medicine

  1999

• postdoctoral fellow, Cell Biology

  University of Virginia

  1994

• Ph.D, Molecular Biology

  National Institute of Immunology

  1991

Awards & honors

• Society for the Study of Reproduction (SSR)
• American Society of Andrology (ASA)
• American Association for the Advancement of Sciences (AAAS)