Investigation of kinesin function and regulation for the purpose of proper chromosome segregation

Mitosis and meiosis are different forms of cell division. Mitosis is a non-reductive form of cell amplification whereby DNA chromosomes are replicated and segregated to form two progeny copies of the progenitor cell. Meiosis is a reductive form of cell division creating progeny containing half the chromosome copies of the progenitor cell. Improper chromosome segregation creates aneuploidy, which is poorly tolerated in cells. In cycling mitotic cells, aneuploidy leads to genome instability and cell death. Following meiosis, aneuploidy is associated with infertility, miscarriages, and birth defects. To segregate chromosome copies properly, pairs are physically organized and segregated to progeny cells by a mitotic spindle, whose functionality is tightly regulated. Kinesins are a family of highly conserved dimeric ATPase proteins which; organize spindle shape and size, facilitate chromosome capture and attachment to the spindle, and generate forces which are required for segregation. I investigated the molecular structure and function of human kinesin 13 family protein, Mitotic Centromere Associated Kinesin, MCAK. MCAK is a microtubule depolymerase whose full molecular structure and mechanism of depolymerization is not fully understood. Using in vitro biochemical assays and in vivo TIRF imaging, I found that altering MCAK molecular structure alters MCAK sub-spindle localization and by inference, alters global microtubule dynamics. This study suggests a potential mode for regulating of MCAK activity/function requiring further testing. Compared to over 30 kinesins in humans, showing a large amount of functional redundancy, yeast only has 6 identified kinesins whose function during meiotic cell division are still relatively unknown. I screened the importance and redundancy of yeast kinesins during meiosis. The results suggest similar roles and redundancies in meiosis to that during mitosis, despite different biochemical and biophysical spindle environments. Together, my investigations broaden the understanding of kinesin regulation and functional redundancy during different types of cell division

Animal microsurgery using microfluidics

Small multicellular genetic organisms form a central part of modern biological research. Using these small organisms provides significant advantages in genetic tractability, manipulation, lifespan and cost. Although the small size is generally advantageous, it can make procedures such as surgeries both time consuming and labor intensive. Over the past few years there have been dramatic improvements in microfluidic technologies that enable significant improvements in microsurgery and interrogation of small multicellular model organisms

Using meta-research methods to examine the inclusion of women, pregnant women, and women-specific health outcomes in studies that contributed to the dietary reference intakes for one-carbon metabolism micronutrients

Purpose: Maternal micronutrient deficiencies occurring during periconceptional, pregnancy, and postpartum periods are a leading cause of adverse pregnancy outcomes globally. The Dietary Reference Intakes (DRIs) are a set of reference values used to assess and guide nutrient intakes of healthy individuals. However, the current DRIs for pregnancy and lactation may be limited in their methods and included populations. The present study analyzed the current DRIs for their inclusion of pregnant women and geographic representativeness. Methods: Meta-research methods were applied to the DRI report for vitamins B6, B12, folate, and choline in four steps: search, screening, full-text data extraction, and data analysis. For each target micronutrient, sections that contributed data to setting the average requirement were focused on, “Selection of Indicators for Estimating the Requirement,” “Findings by Life Stage and Gender Group,” and “Tolerable Upper Intake Limit” for adults, pregnancy, and lactation sub-sections. Screening involved reviewing the reference list to determine whether a reference directly contributed to setting the DRI. Full-text data extraction of primary data was conducted in areas of: 1) administrative information; 2) study methods; 3) human population characteristics; and 4) non-human subjects. Descriptive analyses were performed to describe the inclusion of women, pregnant women, geographic patterns, and demographic diversity. Results: For Vitamin B12, 100% of indicator studies and 71% of life stages studies included women, with a total of 3,246 women participants. However, none of the indicator studies and 15% of life stages studies included pregnant women, with a total of 556 pregnant women participants. None of the indicator studies and 8% of life stages studies reported health measurements specific to women, pregnancy, or lactation. Geographically, 54% of studies were conducted in the United States, and 18% took place in low-and middle-income countries. Data analysis is ongoing for the remaining micronutrients. Conclusions: Preliminary findings indicate that the body of evidence informing the current DRIs are limited in their inclusion of women and pregnant women. Numerous adverse pregnancy and birth outcomes are preventable through optimal maternal nutrition. Therefore, it is critical to ensure that the DRIs are suitable for their intended population. Despite their original intent for use in North America, the DRIs are widely adopted globally. Thus, geographic representation of the studies underlying the DRIs have implications for generalizability

Immune boosting by B.1.1.529 (Omicron) depends on previous SARS-CoV-2 exposure

The Omicron, or Pango lineage B.1.1.529, variant of SARS-CoV-2 carries multiple spike mutations with high transmissibility and partial neutralizing antibody (nAb) escape. Vaccinated individuals show protection from severe disease, often attributed to primed cellular immunity. We investigated T and B cell immunity against B.1.1.529 in triple mRNA vaccinated healthcare workers (HCW) with different SARS-CoV-2 infection histories. B and T cell immunity against previous variants of concern was enhanced in triple vaccinated individuals, but magnitude of T and B cell responses against B.1.1.529 spike protein was reduced. Immune imprinting by infection with the earlier B.1.1.7 (Alpha) variant resulted in less durable binding antibody against B.1.1.529. Previously infection-naïve HCW who became infected during the B.1.1.529 wave showed enhanced immunity against earlier variants, but reduced nAb potency and T cell responses against B.1.1.529 itself. Previous Wuhan Hu-1 infection abrogated T cell recognition and any enhanced cross-reactive neutralizing immunity on infection with B.1.1.529

Quantitative, multiplexed, targeted proteomics for ascertaining variant specific SARS-CoV-2 antibody response

Determining the protection an individual has to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants of concern (VoCs) is crucial for future immune surveillance, vaccine development, and understanding of the changing immune response. We devised an informative assay to current ELISA-based serology using multiplexed, baited, targeted proteomics for direct detection of multiple proteins in the SARS-CoV-2 anti-spike antibody immunocomplex. Serum from individuals collected after infection or first- and second-dose vaccination demonstrates this approach and shows concordance with existing serology and neutralization. Our assays show altered responses of both immunoglobulins and complement to the Alpha (B.1.1.7), Beta (B.1.351), and Delta (B.1.617.1) VoCs and a reduced response to Omicron (B1.1.1529). We were able to identify individuals who had prior infection, and observed that C1q is closely associated with IgG1 (r > 0.82) and may better reflect neutralization to VoCs. Analyzing additional immunoproteins beyond immunoglobulin (Ig) G, provides important information about our understanding of the response to infection and vaccination

Initiation of DNA replication requires actin dynamics and formin activity

International audienc

Limited data exist to inform our basic understanding of micronutrient requirements in pregnancy.

Women and pregnant people have historically been underrepresented in research; this may extend to the basic research informing nutrient reference values, such as the United States’ and Canada’s Dietary Reference Intakes (DRIs). After screening the DRI reports for 23 micronutrients, we extracted metadata from 704 studies. Women were excluded in 23% of studies, and they accounted for a smaller proportion of the sample size (29%). Pregnant or lactating people were included in 17% of the studies. Studies that used rigorous design elements, such as controlled feeding and stable isotope studies, were the most likely to include men only. The majority of studies (>90%) did not report race and ethnicity. Although nutrient reference values are intended for use in the general population, we find that the basic science informing these values may not be generalizable. We call urgently upon funders and researchers to address fundamental gaps in knowledge with high-quality research

Animal microsurgery using microfluidics

Small multicellular genetic organisms form a central part of modern biological research. Using these small organisms provides significant advantages in genetic tractability, manipulation, lifespan and cost. Although the small size is generally advantageous, it can make procedures such as surgeries both time consuming and labor intensive. Over the past few years there have been dramatic improvements in microfluidic technologies that enable significant improvements in microsurgery and interrogation of small multicellular model organisms