Biology subjects

Chiu, K.

Publications and source records attributed to Chiu, K..

6 recordsLinked to original sources

bioRxiv2023

CTCF is essential for proper mitotic spindle structure and anaphase segregation

Mitosis is an essential process in which the duplicated genome is segregated equally into two daughter cells. CTCF has been reported to be present in mitosis but its importance for mitotic fidelity remains to be determined. To evaluate the importance of CTCF in mitosis, we tracked mitotic behaviors in wild type and two different CTCF CRISPR-based genetic knockdowns. We find that knockdown of CTCF results in prolonged mitoses and failed anaphase segregation via time lapse imaging of SiR-DNA. CTCF knockdown did not alter cell cycling or the mitotic checkpoint, which was activated upon nocodazole treatment. Immunofluorescence imaging of the mitotic spindle in CTCF knockdowns revealed disorganization via tri/tetrapolar spindles and chromosomes behind the spindle pole. Imaging of interphase nuclei showed that nuclear size increased drastically, consistent with failure to divide the duplicated genome in anaphase. Population measurements of nuclear shape in CTCF knockdowns do not display decreased circularity or increased nuclear blebbing relative to wild type. However, failed mitoses do display abnormal nuclear morphologies relative to successful mitoses, suggesting population images do not capture individual behaviors. Thus, CTCF is important for both proper metaphase organization and anaphase segregation which impacts the size and shape of the interphase nucleus.

cell biology↗

bioRxiv2022

Actin contraction controls nuclear blebbing and rupture independent of actin confinement

The nucleus is a mechanically stable compartment of the cell that contains the genome and performs many essential functions. Nuclear mechanical components chromatin and lamins maintain nuclear shape, compartmentalization, and function by resisting antagonistic actin contraction and confinement. However, studies have yet to compare chromatin and lamins perturbations side-by-side as well as modulated actin contraction while holding confinement constant. To accomplish this, we used NLS-GFP to measure nuclear shape and rupture in live cells with chromatin decompaction (VPA), loss of lamin B1 (LMNB1-/-), and loss of lamin A/C (LMNA-/-). We then modulated actin contraction while maintaining actin confinement measured by nuclear height. Wild type, chromatin decompaction, and lamin B1 null present bleb-based nuclear deformations and ruptures dependent on actin contraction and independent of actin confinement. Inhibition of actin contraction by Y27632 decreased nuclear blebbing and ruptures to near 0% of cells while activation of actin contraction by CN03 increased the frequency of ruptures by nearly two-fold. However, lamin A/C null results in overall abnormal shape, but similar blebs and ruptures as wild type which were unaffected by actin contraction modulation. Actin contraction control of nuclear shape and ruptures showed that DNA damage levels were more correlated with perturbed nuclear shape than they were with changes in nuclear ruptures. We reveal that lamin B1 is a chromatin decompaction phenotype because using GSK126, which mimics the loss of facultative heterochromatin in lamin B1 null, is sufficient to phenocopy increased nuclear blebbing and ruptures. Furthermore, even though blebs and ruptures in lamin A/C null cells are insensitive to actin contraction, they do have the capacity to form increased levels of nuclear blebs and bleb-based ruptures, shown by treating with VPA. Thus, nuclear bleb formation and bleb-based nuclear ruptures are driven by actin contraction and independent of changes in actin confinement.

cell biology↗

bioRxiv2022

Development and application of genetic ancestry reconstruction methods to study diversity of patient-derived models in the NCI PDXNet Consortium

Personalized medicine holds great promise for improving cancer outcomes, yet there is a large inequity in the demographics of patients from whom genomic data and models, including patient derived xenografts (PDX), are developed and for whom treatments are optimized. In this study we develop a genetic ancestry pipeline for the Cancer Genomics Cloud, which we use to assess the diversity of models currently available in the National Cancer Institute (NCI) supported PDX Development and Trial Centers Research Network (PDXNet). We show that there is an over-representation of models derived from patients of European ancestry, which is consistent with other cancer model resources. We discuss these findings in the context of disparities in cancer incidence and outcomes among demographic groups in the US. For example, for the top cancer health disparities affecting African Americans and Latinos, there is a significant lack of ethnic/race appropriate models needed to advance pre-clinical research and personalized clinical treatment. For stomach and liver tumors, which represent disparities in these two minority populations, there are only three available models derived from patients from such backgrounds. Fortunately, ongoing NCI-funded efforts in minority focused PDXNet centers are actively addressing these gaps. We further discuss these results in the context of power analyses to highlight the immediate need for the development of models from minority populations to address cancer health equity in personalized medicine.

cancer biology↗

bioRxiv2022

Mapping mitonuclear epistasis using a novel recombinant yeast population

Natural genetic variation in mitochondrial and nuclear genomes can influence phenotypes by perturbing coadapted mitonuclear interactions. Mitonuclear epistasis, i.e. non-additive phenotype effects of interacting mitochondrial and nuclear alleles, is emerging as a general feature in eukaryotes, yet very few mitonuclear loci have been identified. Here, we present a novel advanced intercrossed population of S. cerevisiae yeasts, called the Mitonuclear Recombinant Collection (MNRC), designed explicitly for detecting mitonuclear loci contributing to complex traits, and use this population to map the genetic basis to mtDNA loss. In yeast, spontaneous deletions within mtDNAs lead to the petite phenotype that heralded mitochondrial research. We show that in natural populations, rates of petite formation are variable and influenced by genetic variation in nuclear, mtDNAs and mitonuclear interactions. We then mapped nuclear and mitonuclear alleles contributing mtDNA stability using the MNRC by integrating mitonuclear epistasis into a genome-wide association model. We found that associated mitonuclear loci play roles in mitotic growth most likely responding to retrograde signals from mitochondria, while associated nuclear loci with main effects are involved in genome replication. We observed a positive correlation between growth rates and petite frequencies, suggesting a fitness tradeoff between mitotic growth and mtDNA stability. We also found that mtDNA stability was influenced by a mobile mitochondrial GC-cluster that is expanding in certain populations of yeast and that selection for nuclear alleles that stabilize mtDNA may be rapidly occurring. The MNRC provides a powerful tool for identifying mitonuclear interacting loci that will help us to better understand genotype-phenotype relationships and coevolutionary trajectories. Author SummaryGenetic variation in mitochondrial and nuclear genomes can perturb mitonuclear interactions and lead to phenotypic differences between individuals and populations. Despite their importance to most complex traits, it has been difficult to identify the interacting loci. Here, we created a novel population of yeast designed explicitly for mapping mitonuclear loci contributing to complex traits and used this population to map genes influencing the stability of mitochondrial DNA (mtDNA). We found that mitonuclear interacting loci were involved in mitotic growth while non-interacting loci were involved in genome replication. We also found evidence that selection for mitonuclear loci that stabilize mtDNAs occurs rapidly. This work provides insight into mechanisms underlying maintenance of mtDNAs. The mapping population presented here is an important new resource that will help to understand genotype/phenotype relationships and coevolutionary trajectories.

evolutionary biology↗

bioRxiv2020

HP1α is a chromatin crosslinker that controls nuclear and mitotic chromosome mechanics

Chromatin, which consists of DNA and associated proteins, contains genetic information and is a mechanical component of the nucleus. Heterochromatic histone methylation controls nucleus and chromosome stiffness, but the contribution of heterochromatin protein HP1 (CBX5) is unknown. We used a novel HP1 auxin-inducible degron human cell line to rapidly degrade HP1. Degradation did not alter transcription, local chromatin compaction, or histone methylation, but did decrease chromatin stiffness. Single-nucleus micromanipulation reveals that HP1 is essential to chromatin-based mechanics and maintains nuclear morphology, separate from histone methylation. Further experiments with dimerization-deficient HP1I165E indicate that chromatin crosslinking via HP1 dimerization is critical, while polymer simulations demonstrate the importance of chromatin-chromatin crosslinkers in mechanics. In mitotic chromosomes, HP1 similarly bolsters stiffness while aiding in mitotic alignment and faithful segregation. HP1 is therefore a critical chromatin-crosslinking protein that provides mechanical strength to chromosomes and the nucleus throughout the cell cycle and supports cellular functions.

biophysics↗

bioRxiv2018

Mesenchymal stem cells protect retinal ganglion cells from degeneration via mitochondrial donation

Retinal ganglion cell (RGC) degeneration is extremely hard to repair or regenerate and is often coupled with mitochondrial dysfunction. Mesenchymal stem cells (MSCs)-based treatment has been demonstrated beneficial for RGC against degeneration. However, underlying mechanisms of MSC-provided RGC protection are largely unknown other than neuropectective paracrine actions. In this study, we sought to investigate whether mitochondrial donation can preserve RGC functions, in a mitochondrial Ndufs4 deficient mouse model of RGC degeneration. The results revealed intravitreal transplanted by induced pluripotent stem cell derived-MSCs (iPSC-MSC) could donate their mitochondria through crossing inner limited membrane to host RGCs. Furthermore, the donated mitochondria effectively protected against RGC death and largely preserved retinal function in Ndufs4-KO mice. Importantly, the protective effects of mitochondrial donation from MSCs were associated with management of pro-inflammatory cytokines. Our data identified a novel role of MSCs-mitochondrial donation in protection of RGC from degeneration, and highlight a viable therapeutic strategy by manipulating stem cell mitochondrial donation for the treatment of retina degeneration in future.

cell biology↗