Biology subjects

Miller, R. K.

Publications and source records attributed to Miller, R. K..

9 recordsLinked to original sources

bioRxiv2026

Developmental shift in β-catenin localization between nuclear and junctional pools during vertebrate nephron development

Wnt/{beta}-catenin signaling is a critical pathway that regulates nephron progenitor renewal versus differentiation as well as nephron patterning. In addition to its role as a transcription co-factor, {beta}-catenin also functions as a structural component of adherens junctions, where it interacts with cadherins to link cell-cell contacts to the cytoskeleton. However, the relationship between the nuclear and junctional localization of {beta}-catenin during vertebrate nephron development remains poorly understood. To define how endogenous {beta}-catenin localization changes during nephrogenesis, we optimized an accelerated-turnover {beta}-catenin chromobody for live imaging in Xenopus embryos. Using in vivo imaging of Xenopus pronephric development, we visualized endogenous {beta}-catenin within the nuclear, cytoplasmic, and junctional compartments. Across successive developmental stages, {beta}-catenin became progressively enriched at epithelial junctions during nephron maturation while remaining abundant within nuclear and cytoplasmic compartments. Quantitative analyses indicate that epithelial maturation is accompanied by coordinated expansion and partitioning of multiple intracellular {beta}-catenin pools rather than a simple redistribution from nuclear to junctional compartments.

developmental biology↗

bioRxiv2026

Dnmbp interacts with Daam1 to facilitate assembly of cadherin-mediated junctions in epithelializing nephric tubules

The mature kidney contains approximately one million nephrons, and defects arising during nephron development can result in lifelong renal impairment, often culminating in kidney failure and transplantation. Nephric tubule formation requires coordinated epithelial processes, including polarity, adhesion, signaling, and vesicle transport; however, how these processes are integrated during kidney morphogenesis remains unclear. Dynamin binding protein (Dnmbp) is a multi-domain scaffolding protein expressed in human kidneys that is involved in several cellular processes. Using the Xenopus embryonic kidney, we previously demonstrated that Dnmbp is essential for nephrogenesis, yet the mechanisms by which it influences nephron development remain undefined. Here, we identify Dnmbp as a novel interacting partner of the Wnt/planar cell polarity effector Daam1. The interaction between Daam1 and Dnmbp was independently identified in two yeast two-hybrid screens, biochemically verified, and supported by structural modeling predictions of a Daam1-Dnmbp complex. In developing Xenopus laevis kidneys, Dnmbp localized to punctate structures associated with E-cadherin-rich cell-cell contacts. Dnmbp depletion significantly reduced junctional E-cadherin localization in both epithelializing and mature nephric tubules without affecting total E-cadherin levels, indicating a role in E-cadherin recruitment or stabilization at adherens junctions. Furthermore, expression of human DNMBP rescued the junctional defects, confirming the specificity of the loss-of-function phenotype. Together, these findings identify Dnmbp as an essential regulator of kidney development and support a model in which Dnmbp provides a mechanistic link between Wnt/PCP signaling, Cdc42 activation, and adherens junction formation during nephrogenesis.

developmental biology↗

bioRxiv2024

Affective Symptoms in Pregnancy are Associated with the Vaginal Microbiome

Composition of the vaginal microbiome in pregnancy is associated with adverse maternal, obstetric, and child health outcomes. Identifying the sources of individual differences in the vaginal microbiome is therefore of considerable clinical and public health interest. The current study tested the hypothesis that vaginal microbiome composition during pregnancy is associated with an individuals experience of affective symptoms and stress exposure. Data were based on a prospective longitudinal study of a diverse and medically healthy community sample of 275 mother-infant pairs. Affective symptoms and stress exposure and select measures of associated biomarkers (diurnal salivary cortisol, serum measures of sex hormones) were collected at each trimester; self-report, clinical, and medical records were used to collect detailed data on socio-demographic factors and health behavior, including diet and sleep. Vaginal microbiome samples were collected in the third trimester (34-40 weeks) and characterized by 16S rRNA sequencing. Identified taxa were clustered into three community state types (CST1-3) based on dissimilarity of vaginal microbiota composition. Results indicate that depressive symptoms during pregnancy were reliably associated with individual taxa and CST3 in the third trimester. Prediction of functional potential from 16S taxonomy revealed a differential abundance of metabolic pathways in CST1-3 and individual taxa, including biosynthetic pathways for the neuroactive metabolites, serotonin and dopamine. With the exception of bioavailable testosterone, no significant associations were found between symptoms- and stress-related biomarkers and CSTs. Our results provide further evidence of how prenatal psychological distress during pregnancy alters the maternal-fetal microbiome ecosystem that may be important for understanding maternal and child health outcomes. ImportancePrenatal affective symptoms and stress are associated with maternal, obstetric, and child health outcomes, but the mechanisms underlying these links and their application to intervention remain unclear. The findings from this investigation extend prior microbiome-oriented research by demonstrating that the maternal vaginal microbiome composition has a biologically plausible mechanistic link with affective symptoms that also suggest additional clinical applications for assessment and intervention.

microbiology↗

bioRxiv2022

A comparative study of cellular diversity between the Xenopus pronephric and mouse metanephric nephron

The kidney is an essential organ that ensures bodily fluid homeostasis and removes soluble waste products from the organism. The functional units within the kidneys are epithelial tubules called nephrons. These tubules take in filtrate from the blood or coelom and selectively reabsorb nutrients through evolutionarily conserved nephron segments, leaving waste product to be eliminated in the urine. Genes coding for functional transporters are segmentally expressed, enabling nephrons to function as selective filters. The developmental patterning program that generates these segments is of great interest. The Xenopus embryonic kidney, the pronephros, has served as a valuable model to identify genes involved in nephron formation and patterning. Prior work has defined the gene expression profiles of Xenopus epithelial nephron segments via in situ hybridization strategies, but our understanding of the cellular makeup of the Xenopus pronephric kidney remains incomplete. Here, we scrutinize the cellular composition of the Xenopus pronephric nephron through comparative analyses with previous Xenopus studies and single-cell mRNA sequencing of the adult mouse kidney, this study reconstructs the cellular makeup of the pronephric kidney and identifies conserved cells, segments, and expression profiles. The data highlight significant conservation in podocytes, proximal and distal tubule cells and divergence in cellular composition underlying the evolution of the corticomedullary axis, while emphasizing the Xenopus pronephros as a model for physiology and disease. Summary StatementAssessment of the conservation of gene expression between functional Xenopus laevis pronephric and mouse metanephric nephrons through single-cell RNA sequencing.

evolutionary biology↗

bioRxiv2022

The TOG protein Stu2 is regulated by acetylation

Stu2 in S. cerevisiae is a member of the XMAP215/Dis1/Alp14/Msps/CKAP5/ch-TOG family of MAPs and has multiple functions in controlling microtubules, including microtubule polymerization, microtubule depolymerization, linking chromosomes to the kinetochore, and assembly of {gamma}-TuSCs at the SPB. Whereas phosphorylation has been shown to be critical for Stu2 localization at the kinetochore, other regulatory mechanisms that control Stu2 function are still poorly understood. Here, we show that a novel form of Stu2 regulation occurs through the acetylation of three lysine residues at K252, K469, and K870, which are located in three distinct domains of Stu2. Alteration of acetylation through acetyl-mimetic and acetyl-blocking mutations did not impact the essential function of Stu2. Instead, these mutations lead to both positive and negative changes in chromosome stability, as well as changes in resistance to the microtubule depolymerization drug, benomyl. In agreement with our in silico modeling, several acetylation-mimetic mutants displayed increased interactions with {gamma}-tubulin. Taken together, these data suggest that Stu2 acetylation can govern multiple Stu2 functions in both a positive and negative manner, including chromosome stability and interactions at the SPB.

genetics↗

bioRxiv2020

Cell-Cell Adhesion During Nephron Development Is Driven by Wnt/PCP Formin Daam1

E-cadherin junctions facilitate the assembly and disassembly of cell-cell contacts that drive development and homeostasis of epithelial tissues. The stability of E-cadherin-based junctions highly depends on their attachment to the actin cytoskeleton, but little is known about how the assembly of junctional actin filaments is regulated. Formins are a conserved group of proteins responsible for the formation and elongation of filamentous actin (F-actin). In this study, using Xenopus embryonic kidney and Madin-Darby canine kidney (MDCK) cells, we investigate the role of the Wnt/ planar cell polarity (PCP) formin protein Daam1 (Dishevelled-associated activator of morphogenesis 1) in regulating E-cadherin based intercellular adhesion. Using live imaging we show that Daam1 localizes to newly formed cell-cell contacts in the developing nephron. Furthermore, analyses of junctional F-actin upon Daam1 depletion indicate a decrease in microfilament localization and their slowed turnover. We also show that Daam1 is necessary for efficient and timely localization of junctional E-cadherin, which is mediated by Daam1s formin homology domain 2 (FH2). Finally, we establish that Daam1 signaling is essential for promoting organized movement of renal cells. This study demonstrates that Daam1 formin junctional activity is critical for epithelial tissue organization.

developmental biology↗

bioRxiv2019

DYRK1A-related intellectual disability: a syndrome associated with congenital anomalies of the kidney and urinary tract

PurposeHaploinsufficiency of DYRK1A causes a recognizable clinical syndrome. The goal of this paper is to investigate congenital anomalies of the kidney and urinary tract (CAKUT) and genital defects (GD) in patients with DYRK1A mutations.\n\nMethodsA large database of clinical exome sequencing (ES) was queried for de novo DYRK1A mutations and CAKUT/GD phenotypes were characterized. Xenopus laevis (frog) was chosen as a model organism to assess Dyrk1as role in renal development.\n\nResultsPhenotypic details and mutations of 19 patients were compiled after an initial observation that one patient with a de novo pathogenic mutation in DYRK1A had GD. CAKUT/GD data were available from 15 patients, 11 of whom present with CAKUT/GD. Studies in Xenopus embryos demonstrate that knockdown of Dyrk1a disrupts the development of segments of developing embryonic nephrons, which ultimately give rise to the entire genitourinary (GU) tract. These defects could be rescued by co-injecting wildtype human DYRK1A RNA, but not with truncated DYRK1AR205* RNA.\n\nConclusionEvidence supports routine GU screening of all individuals with de novo DYRK1A pathogenic variants to ensure optimized clinical management. Collectively, the reported clinical data and loss of function studies in Xenopus substantiate a novel role for DYRK1A in GU development.

genetics↗

bioRxiv2019

The Role of the Wnt/PCP Formin Daam1 in Renal Ciliogenesis

Kidneys are composed of numerous ciliated epithelial tubules called nephrons. Each nephron functions to reabsorb nutrients and concentrate waste products into urine. Defects in primary cilia are associated with abnormal formation of nephrons and cyst formation in a wide range of kidney disorders. Previous work in Xenopus laevis and zebrafish embryos established that loss of components that make up the Wnt/PCP pathway, Daam1 and ArhGEF19 (wGEF) perturb kidney tubulogenesis. Dishevelled, which activates both the canonical and non-canonical Wnt/PCP pathway, affect cilia formation in multiciliated cells. In this study, we investigated the role of the noncanoncial Wnt/PCP components Daam1 and ArhGEF19 (wGEF) in renal ciliogenesis utilizing polarized mammalian kidney epithelia cells (MDCKII and IMCD3) and Xenopus laevis embryonic kidney. We demonstrate that knockdown of Daam1 and ArhGEF19 in MDCKII and IMCD3 cells leads to loss of cilia, and Daam1s effect on ciliogenesis is mediated by the formin-activity of Daam1. Moreover, Daam1 co-localizes with the ciliary transport protein IFT88. Interestingly, knocking down Daam1 in Xenopus kidney does not lead to loss of cilia. This data suggests a new role for Daam1 in the formation of primary cilia.

molecular biology↗

bioRxiv2018

Dynamin binding protein is required for Xenopus laevis kidney development

The adult human kidney contains over one million nephrons, with each nephron consisting of a tube containing segments that have specialized functions in nutrient and water absorption and waste excretion. The embryonic kidney of Xenopus laevis consists of a single functional nephron composed of regions that are analogous to those found in the human nephron, making it a simple model for the study of nephrogenesis. The exocyst complex, which traffics proteins to the cell membrane in vesicles via CDC42, is essential for normal kidney development. Here, we show that the CDC42-GEF, dynamin binding protein (Dnmbp/Tuba), is essential for nephrogenesis in Xenopus. dnmbp is expressed in Xenopus embryo kidneys during development, and knockdown of Dnmbp using two separate morpholino antisense oligonucleotides results in reduced expression of late pronephric markers, whereas the expression of early markers of nephrogenesis remains unchanged. A greater reduction in expression of markers of differentiated distal and connecting tubules was seen in comparison to proximal tubule markers, indicating that Dnmbp reduction may have a greater impact on distal and connecting tubule differentiation. dnmbp knockout using CRISPR results in a similar reduction of late markers of pronephric tubulogenesis. Overexpression of dnmbp in the kidney also resulted in disrupted pronephric tubules, suggesting that dnmbp levels in the developing kidney are tightly regulated, with either increased or decreased levels leading to developmental defects. Together, these data suggest that Dnmbp is required for nephrogenesis.

developmental biology↗