Biology subjects

Brieno-Enriquez, M. A.

Publications and source records attributed to Brieno-Enriquez, M. A..

6 recordsLinked to original sources

bioRxiv2026

Tex11 Mutant Mouse Models of Human Azoospermia

Non-obstructive azoospermia (NOA) is the absence of sperm in the ejaculate due to spermatogenic failure. Fifty percent of NOA cases are unexplained but may arise from unidentified genetic mutations. Variants in TEX11 have been identified in men with NOA; and Tex11 knockout in mice causes NOA. Here we attempt to validate three TEX11 variants discovered in NOA patients by knocking them into the orthologous region of the mouse genome using CRISPR/Cas9 gene editing. Compared to wild type (144.2 {+/-} 9.87 mg; 1.7 {+/-} 0.5 million sperm/cauda epididymis 4.8 {+/-} 1.3 pups/breeding), Tex11D mice (frameshift mutation) had reduced testis weight (28.33 {+/-} 1.16 mg); no sperm in the epididymis; and were infertile with a maturation arrest testicular phenotype. We did not observe any spermatogenesis or fertility defects Tex11A mice (missense mutation). Tex11L mice had reduced testis weight (87.5 {+/-} 14.79 mg) and epididymal sperm counts (0.33{+/-}0.13 million/cauda epididymis) but an incompletely penetrant infertility phenotype (5.4 {+/-} 1.13 pups/breeding) with one third of mice being infertile. Infertile Tex11L mice also had a distinct epididymal phenotype with reduced sperm density in the caput and no sperm in the cauda, which was filled with amorphous material.

genetics↗

bioRxiv2024

EXOSOMES FROM CYCLIC MICE MODULATE LIVER TRANSCRIPTOME IN ESTROUPAUSE MICE INDEPENDENT OF AGE

BackgroundExosomes are extracellular vesicles secreted by cells that contain microRNAs (miRNAs). These miRNAs can induce changes in gene expression and function of recipient cells. In different cells exosome content can change with age and physiological state affecting tissues function and health. AimsTherefore, the aim of this study was to characterize the miRNA content and role of exosomes from cyclic female mice in the modulation of liver transcriptome in estropausal mice. Main MethodsTwo-month-old female mice were induced to estropause using 4-vinylcyclohexene diepoxide (VCD). At six months of age VCD-treated mice were divided in control group (VCD) and exosome treated group (VCD+EXO), which received 10 injections at 3-day intervals of exosomes extracted from serum of cyclic female mice (CTL). Key findingsExosome injection in estropausal mice had no effect on body mass, insulin sensitivity or organ weight. We observed ten miRNAs differentially regulated in serum exosomes of VCD compared to CTL mice. In the liver we observed 931 genes differentially expressed in VCD+EXO compared to VCD mice. Interestingly, eight pathways were up-regulated in liver by VCD treatment and down-regulated by exosome treatment, indicating that exosomes from cyclic mice can reverse changes promoted by estropause in liver. Cyp4a12a expression which is male-specific was increased in VCD females and not reversed by exosome treatment. SignificanceOur findings indicate that miRNAs content in exosomes is regulated by estropause in mice independent of age. Additionally, treatment of estropausal mice with exosomes from cyclic mice can partially reverse changes in liver transcriptome.

physiology↗

bioRxiv2021

Multiple 9-1-1 complexes promote homolog synapsis, DSB repair, and ATR signaling during mammalian meiosis

DNA damage response mechanisms have meiotic roles that ensure successful gamete formation. While completion of meiotic double-strand break (DSB) repair requires the canonical RAD9A-RAD1-HUS1 (9A-1-1) complex, mammalian meiocytes also express RAD9A and HUS1 paralogs, RAD9B and HUS1B, predicted to form alternative 9-1-1 complexes. The RAD1 subunit is shared by all predicted 9-1-1 complexes and localizes to meiotic chromosomes even in the absence of HUS1 and RAD9A. Here we report that testis-specific RAD1 disruption resulted in impaired DSB repair, germ cell depletion and infertility. Unlike Hus1 or Rad9a disruption, Rad1 loss also caused defects in homolog synapsis, ATR signaling and meiotic sex chromosome inactivation. Comprehensive testis phosphoproteomics revealed that RAD1 and ATR coordinately regulate numerous proteins involved in DSB repair, meiotic silencing, synaptonemal complex formation, and cohesion. Together, these results establish critical roles for both canonical and alternative 9-1-1 complexes in meiotic ATR activation and successful prophase I completion.

genetics↗

bioRxiv2019

Mutation Of MLH3 Endonuclease Motif Reveals Integration Between Crossover Pathways In Mammalian Meiosis

During meiotic prophase I, double-strand breaks (DSBs) initiate homologous recombination leading to non-crossovers (NCOs) and crossovers (COs). In mouse, 10% of DSBs are designated to become COs, primarily through a pathway dependent on the MLH1-MLH3 heterodimer (MutL{gamma}). Mlh3 contains an endonuclease domain that is critical for resolving COs in yeast. We generated a mouse (Mlh3DN/DN) harboring a mutation within this conserved domain that is predicted to generate a protein that is catalytically inert. Mlh3DN/DN males, like fully null Mlh3-/- males, have no spermatozoa and are infertile, yet spermatocytes have normal DSBs and undergo normal synapsis events in early prophase I. Unlike Mlh3-/- males, mutation of the endonuclease domain within MLH3 permits normal loading and frequency of MutL{gamma} in pachynema. However, key DSB repair factors (RAD51) and mediators of CO pathway choice (BLM helicase) persist into pachynema in Mlh3DN/DN males, indicating a temporal delay in repair events and revealing a mechanism by which alternative DSB repair pathways may be selected. While Mlh3DN/DN spermatocytes retain only 22% of wildtype chiasmata counts, this frequency is greater than observed in Mlh3-/- males (10%), suggesting that the allele may permit partial endonuclease activity, or that other pathways can generate COs from these MutL{gamma}-defined repair intermediates in Mlh3DN/DN males. Double mutant mice homozygous for the Mlh3DN/DN and Mus81-/- mutations show losses in chiasmata that approach levels observed in Mlh3-/- males, indicating that the MUS81-EME1-regulated crossover pathway accounts for some of the increased residual chiasmata observed in Mlh3DN/DN spermatocytes. Our data demonstrate that mouse spermatocytes bearing the MLH1-MLH3DN/DN complex display the proper loading of factors essential for CO resolution (MutS{gamma}, CDK2, HEI10, MutL{gamma}). Despite these functions, mice bearing the Mlh3DN/DN allele show defects in the repair of meiotic recombination intermediates and a loss of most chiasmata. SUMMARYThe MLH1-MLH3 complex is essential for crossing over in mammalian meiosis. We generated a mutation in mouse MLH3 that alters its conserved endonuclease domain and show that it disrupts crossing over in a manner distinct from the full null Mlh3 mouse, but also results in male infertility.

genetics↗

bioRxiv2018

Characterization of Telomeric Repeat-Containing RNA (TERRA) localization and protein interactions in Primordial Germ Cells of the mouse

Telomeres are dynamic nucleoprotein structures capping the physical ends of linear eukaryotic chromosomes. They consist of telomeric DNA repeats (TTAGGG), the shelterin protein complex, and Telomeric Repeat-Containing RNA (TERRA). Proposed TERRA functions are wide-ranging and include telomere maintenance, telomerase inhibition, genomic stability, and alternative lengthening of telomere. However, the role of TERRA in primordial germ cells (PGCs), the embryonic precursors of germ cells, is unknown. Using RNA-fluorescence in situ hybridization (RNA-FISH) we identify TERRA in PGCs soon after these cells have migrated to, and become established in, the developing gonad. RNA-FISH showed the presence of TERRA transcripts in female PGCs at 11.5, 12.5 and 13.5 days post-coitum. In male PGCs, however, TERRA transcripts are observable from 12.5 dpc. Using qPCR we evaluated chromosome-specific TERRA expression, and demonstrated that TERRA levels vary with sex and gestational age, and that transcription of TERRA from specific chromosomes is sexually dimorphic. TERRA interacting proteins were evaluated using Identification of Direct RNA Interacting Proteins (iDRiP) which identified 48 in female and 26 in male protein interactors specifically within nuclear extracts from PGCs at 13.5 dpc. We validated two different proteins the splicing factor, proline- and glutamine-rich (SFPQ) in PGCs and Non-POU domain-containing octamer-binding protein (NONO) in somatic cells. Our results show that, TERRA interacting proteins are determined by sex in both PGCs and somatic cells. Taken together, our data indicate that TERRA expression and interactome during PGC development are regulated in a dynamic fashion that is dependent on gestational age and sex.\n\nResearch reported in this publication was supported in part by the Eunice Kennedy Shriver National Institute of Child Health & Human Development of the National Institutes of Health under Award Number K99HD090289 to M.A.B-E. A seed grant from the Cornell Center for Reproductive Genomics to M.A.B-E, using funds obtained as part of the NICHD National Centers for Translational Research in Reproduction and Infertility (NCTRI), award number P50HD076210 to P.E.C. and Empire State Stem Cell Fund through New York State Department of Health Contract # C30293GG. Imaging data was acquired through the Cornell University Biotechnology Resource Center, with NSF funding #1428922 for the shared Zeiss Elyra Microscope. NIH SIG 1S10 OD017992-01 grant support the Orbitrap Fusion mass spectrometer. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

genetics↗

bioRxiv2017

NIMA-related kinase 1 (NEK1) regulates the localization and phosphorylation of α-Adducin (ADD1) and Myosin X (MYO10) during meiosis

Summary statementNEK1 kinase regulates the assembly and function of the meiosis I spindle by phosphorylating -adducin (ADD1) and thereby facilitating its interaction with Myosin X (MYO10)\n\nAbstractNIMA-related kinase 1 (NEK1) is a serine/threonine and tyrosine kinase that is highly expressed in mammalian germ cells. Mutations in Nek1 induce anemia, polycystic kidney and infertility. In this study we evaluated the role of NEK1 in meiotic spindle formation in both male and female gametes. Our results show that the lack of NEK1 provokes an abnormal organization of the meiosis I spindle characterized by elongated and/or multipolar spindles, and abnormal chromosome congression. The aberrant spindle structure is concomitant with the disruption in localization and protein levels of myosin X (MYO10) and -adducin (ADD1), both of which are implicated in the regulation of spindle formation during mitosis. Interaction of ADD1 with MYO10 is dependent on phosphorylation, whereby phosphorylation of ADD1 enables its binding to MYO10 on mitotic spindles. Reduction in ADD1 protein in NEK1 mutant mice is associated with hyperphosphorylation of ADD1, thereby preventing the interaction with MYO10 during meiotic spindle formation. Our results reveal a novel regulatory role for NEK1 in the regulation of spindle architecture and function during meiosis.

cell biology↗