Biology subjects

Hannum, D. F.

Publications and source records attributed to Hannum, D. F..

5 recordsLinked to original sources

bioRxiv2025

Transcriptomic and proteomic dynamics of ovarian follicle group culture resemble in vivo folliculogenesis

The prohibitively low yield of fertilizable oocytes obtained from cultured ovarian follicles limits clinical translation of in vitro follicle maturation for fertility preservation. This is in part due to an incomplete understanding of the process of follicle development. Previous work has demonstrated that group culture of primary murine follicles had a synergistic effect on growth and maturation in contrast to single follicles, but mechanisms remained unknown. Here, we cultured primary follicles in groups of 5 (5X) or 10 (10x) for twelve days, separated the somatic cells from oocytes, and analyzed the temporal transcriptional signatures every two days. In total, 13,461 genes in somatic cells and 10,091 genes in oocytes were computationally sorted into ten temporally distinct gene expression patterns. The somatic cell temporal gene expression patterns showed strong concordance with the granulosa and theca cell markers reported in a recent single-cell whole ovary RNA sequencing study. Importantly, canonical markers of steroidogenesis in cultured follicles followed expected trajectories of decreasing Amh expression and increasing Inhba, Inhbb, Cyp11a1, Cyp17a1, Cyp19a1, Lhcgr, and Fshr over the culture period. Furthermore, when comparing the 10X and 5X culture groups, we identified 306 and 14 differentially expressed genes in somatic cells and oocytes, respectively. Shotgun proteomics data was aligned with the somatic cell transcriptomic data and identified four L-R pairs that were differentially expressed between the two conditions. These comprehensive datasets uncovered temporal dynamics of in vitro folliculogenesis in a compartment-specific manner, serving as a valuable resource for optimizing future follicle culture systems for fertility preservation.

cell biology↗

bioRxiv2025

Single-oocyte transcriptional profile of early-stage human oocytes reveals differentially expressed genes in the primordial and transitioning stages.

The critical initial step in human oocyte maturation - the transition of ovarian follicles from dormancy to activation - remains poorly understood. Here we performed RNA sequencing on single oocytes isolated from early-stage follicles from nine healthy reproductive-age donors. Data for 133 high-quality oocytes formed two connected clusters, C1 and C2, with 5,449 significantly differentially expressed genes. Using recently reported gene lists for early-stage follicles we found that C1 oocytes likely came from earlier, dormant primordial follicles, while C2 oocytes match later-stage primordial or transitioning follicles. We sought to validate two DE genes, UHRF1 for C1 and CCN2 for C2, by their RNA-FISH in situ pattern in morphologically classified follicles, but did not observe statistically significant differences between follicle stages. This apparent discrepancy between follicles stage determined by its morphology and oocytes transcriptional state, if replicated in additional studies, may indicate a lack of closely coupled synchrony between follicle morphology and oocytes functional state.

cell biology↗

bioRxiv2025

Splicing variants in MYRF cause partial loss of function in the retinal pigment epithelium

Myelin Regulatory Factor (MYRF) regulates retinal pigment epithelial (RPE) development and variants in the C-terminus are linked to isolated nanophthalmos, while loss-of-function variants cause syndromic disease. To define the molecular mechanism of this discrepancy, in vitro and animal studies were performed on a pathogenic C-terminal variant (p.Gly1126fs30* or dG-MYRF). ARPE-19 cells transduced with dG-MYRF revealed reduced target gene expression compared to WT-MYRF, with reduced steady state levels of C-terminal MYRF cleavage product, but intact cleavage and localization. A homozygous humanized MYRF C-terminal (MyrfhumdG/humdG) mouse model was embryonic lethal by embryonic day (E) 18.5, while humanized wildtype (MyrfhumWT/humWT) showed normal expression and survival. Bioinformatic analysis on integrated single cell RNA-seq from humanized E17.5 and knockout Rx-Cre;Myrffl/fl (E15.5 and P0) mice supported shared differentially expressed genes with decreased effect size in MyrfhumdG/humdG eyes. These findings, and the viability differences, support that dG-MYRF is a hypomorphic allele. Further, two novel MYRF splicing variants were identified in families with isolated nanophthalmos, with one confirmed to alter 40% of spliced transcripts, creating a nonfunctional isoform. These cases corroborate that isolated nanophthalmos results from hypomorphic alleles of MYRF, supporting a tissue-specific threshold effect and suggests that the C-terminus has unique roles in the RPE.

genetics↗

bioRxiv2024

Cellular heterogeneity and dynamics of the human uterus in healthy premenopausal women

The human uterus is a complex and dynamic organ whose lining grows, remodels, and regenerates in every menstrual cycle or upon tissue damage. Here we applied single-cell RNA sequencing to profile more the 50,000 uterine cells from both the endometrium and myometrium of 5 healthy premenopausal individuals, and jointly analyzed the data with a previously published dataset from 15 subjects. The resulting normal uterus cell atlas contains more than 167K cells representing the lymphatic endothelium, blood endothelium, stromal, ciliated epithelium, unciliated epithelium, and immune cell populations. Focused analyses within each major cell type and comparisons with subtype labels from prior studies allowed us to document supporting evidence, resolve naming conflicts, and to propose a consensus annotation system of 39 subtypes. We release their gene expression centroids, differentially expressed genes, and mRNA patterns of literature-based markers as a shared community resource. We find many subtypes show dynamic changes over different phases of the cycle and identify multiple potential progenitor cells: compartment-wide progenitors for each major cell type, transitional cells that are upstream of other subtypes, and potential cross-lineage multipotent stromal progenitors that may be capable of replenishing the epithelial, stromal, and endothelial compartments. When compared to the healthy premenopausal samples, a postpartum and a postmenopausal uterus sample revealed substantially altered tissue composition, involving the rise or fall of stromal, endothelial, and immune cells. The cell taxonomy and molecular markers we report here are expected to inform studies of both basic biology of uterine function and its disorders. SIGNIFICANCEWe present single-cell RNA sequencing data from seven individuals (five healthy pre-menopausal women, one post-menopausal woman, and one postpartum) and perform an integrated analysis of this data alongside 15 previously published scRNA-seq datasets. We identified 39 distinct cell subtypes across four major cell types in the uterus. By using RNA velocity analysis and centroid-centroid comparisons we identify multiple computationally predicted progenitor populations for each of the major cell compartments, as well as potential cross-compartment, multi-potent progenitors. While the function and interactions of these cell populations remain to be validated through future experiments, the markers and their "dual characteristics" that we describe will serve as a rich resource to the scientific community. Importantly, we address a significant challenge in the field: reconciling multiple uterine cell taxonomies being proposed. To achieve this, we focused on integrating historical and contemporary knowledge across multiple studies. By providing detailed evidence used for cell classification we lay the groundwork for establishing a stable, consensus cell atlas of the human uterus.

cell biology↗

bioRxiv2021

Cellular heterogeneity of human fallopian tubes in normal and hydrosalpinx disease states identified by scRNA-seq

Fallopian tube (FT) homeostasis requires dynamic regulation of heterogeneous cell populations and is disrupted in infertility and ovarian cancer. Here we applied single-cell RNAseq to profile 53,376 FT cells from 3 healthy pre-menopausal subjects. The resulting cell atlas contains 12 major cell types representing epithelial, stromal and immune compartments. Re-clustering of epithelial cells identified 4 ciliated and 6 non-ciliated secretory epithelial subtypes, two of which represent potential progenitor pools: one leading to mature secretory cells, while the other contributing to either ciliated cells or one of the stromal cell types. To understand how FT cell numbers and states change in a disease state, we analyzed ~15,000 cells from a hydrosalpinx sample and observed shifts in epithelial and stromal populations, and cell type-specific changes in extracellular matrix and TGF-{beta} signaling, underscoring fibrosis pathophysiology. This resource is expected to facilitate future studies to understand fallopian tube homeostasis in normal development and disease.

systems biology↗