Biology subjects

Gonzalez, L. E.

Publications and source records attributed to Gonzalez, L. E..

7 recordsLinked to original sources

bioRxiv2023

Distribution and dynamics of chromatin states in the C. elegans germ line

Chromatin organization in the C. elegans germ line is tightly regulated and critical for germ cell differentiation. While certain germline epigenetic regulatory mechanisms have been identified, how they influence chromatin structure and ultimately gene expression remain unclear, in part because most genomic studies have focused on data collected from whole worms. We therefore analyzed publicly available histone modification and chromatin accessibility data from isolated undifferentiated germ nuclei to define chromatin states. We then correlated these states with overall transcript abundance, spatio-temporal expression patterns, and the function of small RNA pathways. Because the essential role of the germ line is to transmit genetic information to the next generation and establish gene expression in the early embryo, we compared epigenetic and transcriptomic profiles from undifferentiated germ cells, oocytes, and embryos to define the epigenetic changes during this developmental transition. The active histone modification H3K4me3 exhibits particularly dynamic remodeling as germ cells differentiate into oocytes. Our results highlight the dynamism of the chromatin landscape in germ cells, and provide a resource for future investigation into epigenetic regulatory mechanisms.

genomics↗

bioRxiv2023

The Upstream Sequence Transcription Complex Dictates Nucleosome Positioning and Promoter Accessibility at piRNA Genes in the C. elegans Germ Line

The piRNA pathway is a conserved germline-specific small RNA pathway that ensures genomic integrity and continued fertility. In C. elegans and other nematodes, Type-I piRNA precursor transcripts are expressed from over 10,000 small, independently regulated genes clustered within two discrete domains of 1.5 and 3.5 MB on Chromosome IV. These large clusters likely play a significant role in promoting germline-specific expression of piRNAs, but the underlying mechanisms are unclear. By examining the chromatin environment specifically in isolated germ nuclei, we demonstrate that piRNA clusters are located in closed chromatin, and confirm the enrichment for the inactive histone modification H3K27me3. We further show that the piRNA biogenesis factor USTC (Upstream Sequence Transcription Complex) plays two roles - it promotes a strong association of nucleosomes throughout the piRNA clusters, and it organizes the local nucleosome environment to direct the exposure of individual piRNA genes. Overall, this work reveals new insight into how chromatin state coordinates transcriptional regulation over large genomic domains, which has implications for understanding global genome organization in the germ line.

genomics↗

bioRxiv2022

Differential metabolic adaptations define responses of winner and loser oncogenic mutant stem cells in skin epidermis in vivo

Skin epithelial stem cells detect and correct aberrancies induced by oncogenic mutations. Different oncogenes invoke different mechanisms of epithelial tolerance: while wild-type cells outcompete {beta}-catenin-Gain-of-Function ({beta}catGOF) mutant cells, HrasG12V mutant cells outcompete wild-type cells1,2. Here we ask how metabolic states change as wild-type stem cells interface with mutant cells, and how this ultimately drives different cell competition outcomes. By adapting our live-imaging platform to track endogenous redox ratio (NAD(P)H/FAD) with single cell resolution in the same mice over time, we show that wild-type epidermal stem cells maintain robust redox ratio despite their heterogeneous cell cycle states. We discover that both {beta}catGOF and HrasG12V models lead to a rapid drop in redox ratios. However, the "winner" cells in each model (wild-type in {beta}catGOF and mutant in HrasG12V), rapidly recover their redox ratios, irrespective of the mutation induced. Using mass spectrometry (13C-LC-MS/MS)3, we find that both mutants increase flux through the oxidative tricarboxylic acid cycle, but the "winner" HrasG12V cells and the "loser" {beta}catGOF cells modulate glycolytic flux differently. Hence, we reveal the metabolic adaptations that define the hallmarks of winners and losers during cell competition in vivo and uncover the nodes of regulation unique to each cell fate.

cancer biology↗

bioRxiv2022

Live imaging reveals chromatin compaction transitions and dynamic transcriptional bursting during stem cell differentiation in vivo

Stem cell differentiation requires dramatic changes in gene expression and global remodeling of chromatin architecture. How and when chromatin remodels relative to the transcriptional, behavioral, and morphological changes during differentiation remain unclear, particularly in an intact tissue context. Here, we develop a quantitative pipeline which leverages fluorescently-tagged histones and longitudinal imaging to track large-scale chromatin compaction changes within individual cells in a live mouse. Applying this pipeline to epidermal stem cells, we reveal that cell-to-cell chromatin compaction heterogeneity within the stem cell compartment emerges independent of cell cycle status, and instead is reflective of differentiation status. Chromatin compaction state gradually transitions over days as differentiating cells exit the stem cell compartment. Moreover, establishing live imaging of keratin-10 nascent RNA, which marks the onset of stem cell differentiation, we find that keratin-10 transcription is highly dynamic and largely precedes the global chromatin compaction changes associated with differentiation. Together, these analyses reveal that stem cell differentiation involves dynamic transcriptional states and gradual chromatin rearrangement.

genetics↗

bioRxiv2022

Oncogenic Kras induces spatiotemporally specific tissue deformation through converting pulsatile into sustained ERK activation

Tissue regeneration and maintenance rely on coordinated stem cell behaviors. This orchestration can be impaired by oncogenic mutations leading to tissue architecture disruption and ultimately cancer formation. However, it is still largely unclear how oncogenes perturb stem cells functions to break tissue architecture. Here, we used intravital imaging and a novel signaling reporter to investigate the mechanisms by which oncogenic Kras mutation causes tissue disruption in the hair follicle. Through longitudinally tracking the same hair follicles in live mice, we found that KrasG12D, a mutation that can lead to squamous cell carcinoma, induces epithelial tissue deformation in a spatiotemporally specific manner. This tissue architecture abnormality is linked with a spatial dysregulation of stem cell proliferation as well as abnormal migration during hair follicle growth. By using a reporter mouse that allows us to capture real-time ERK signal dynamics at the single cell level, we discovered that KrasG12D, but not a closely related mutation HrasG12V, converts the pulsatile ERK signal fluctuation in the stem cells into sustained activation. Furthermore, by combining drug treatment with longitudinal imaging, we demonstrated that temporary inhibiting ERK signal reverts the KrasG12D-induced tissue deformation, suggesting that sustained ERK activation leads to tissue architecture disruption in Kras mutant hair follicles. Altogether, our work suggests that oncogenic mutations induce tissue abnormalities when spatiotemporally specific conditions are met, which allows mutant stem cells to disturb local cell coordination through altering dynamic signal communications.

cancer biology↗

bioRxiv2022

Reorganizing Niche Architecture Still Preserves Organ Function in the Hair Follicle

Stem cells ability to build and replenish tissues depends on support from their niche. While niche architecture varies across different organs, the functional importance of niche architecture is unclear. During hair follicle growth, multipotent epithelial progenitors build hair via crosstalk with their remodeling fibroblast niche, the dermal papilla, providing a powerful model to functionally interrogate different niche architectures. Through intravital imaging, we show that dermal papilla fibroblasts remodel both individually and collectively to form a polarized, structurally robust niche. Polarized TGF{beta} signaling precedes structural niche polarity, and loss of TGF{beta} signaling in dermal papilla fibroblasts leads them to progressively lose their stereotypic architecture and instead surround the epithelium. The reorganized niche relocates multipotent progenitors, but nevertheless supports their proliferation and differentiation. However, progenitor differentiation is completed prematurely, resulting in compromised hair production. Overall, our results reveal that niche architecture optimizes organ efficiency, but is not absolutely essential for organ function.

developmental biology↗

bioRxiv2021

Maternal Piwi Regulates Primordial Germ Cell Development to Ensure the Fertility of Female Progeny in Drosophila

In many animals, germline development is initiated by proteins and RNAs that are expressed maternally. PIWI proteins and their associated small noncoding PIWI-interacting RNAs (piRNAs), which guide PIWI to target RNAs by base-pairing, are among the maternal components deposited into the germline of the early embryo in Drosophila. Piwi has been extensively studied in the adult ovary and testis, where it is required for transposon suppression, germline stem cell self-renewal, and fertility. Consequently, loss of Piwi in the adult ovary using piwi-null alleles or knockdown from early oogenesis results in complete sterility, limiting investigation into possible embryonic functions of maternal Piwi. In this study, we show that the maternal Piwi protein persists in the embryonic germline through gonad coalescence, suggesting that maternal Piwi can regulate germline development beyond early embryogenesis. Using a maternal knockdown strategy, we find that maternal Piwi is required for the fertility and normal gonad morphology of female, but not male, progeny. Following maternal piwi knockdown, transposons were mildly derepressed in the early embryo but were fully repressed in the ovaries of adult progeny. Furthermore, the maternal piRNA pool was diminished, reducing the capacity of the PIWI/piRNA complex to target zygotic genes during embryogenesis. Examination of embryonic germ cell proliferation and ovarian gene expression showed that the germline of female progeny was partially masculinized by maternal piwi knockdown. Our study reveals a novel role for maternal Piwi in the germline development of female progeny and suggests that the PIWI/piRNA pathway is involved in germline sex determination in Drosophila.

genetics↗