Holly Gibbs - Academia.edu (original) (raw)

Papers by Holly Gibbs

Development Genes and Evolution, Mar 1, 2019

One of the earliest patterning events in the vertebrate neural plate is the specification of mes/... more One of the earliest patterning events in the vertebrate neural plate is the specification of mes/r1, the territory comprising the prospective mesencephalon and the first hindbrain rhombomere. Within mes/r1, an interface of gene expression defines the midbrain-hindbrain boundary (MHB), a lineage restriction that separates the mesencephalon and rhombencephalon. wnt1 is critical to mes/r1 development and functions within the MHB as a component of the MHB gene regulatory network (GRN). Despite its importance to these critical and early steps of vertebrate neurogenesis, little is known about the factors responsible for wnt1 transcriptional regulation. In the zebrafish, wnt1 and its neighboring paralog, wnt10b, are expressed in largely overlapping patterns, suggesting co-regulation. To understand wnt1 and wnt10b transcriptional control, we used a comparative genomics approach to identify relevant enhancers. We show that the wnt1-wnt10b locus contains multiple cis-regulatory elements that likely interact to generate the wnt1 and wnt10b expression patterns. Two of eleven conserved enhancers tested show activity restricted to the midbrain and MHB, an activity that is conserved in the distantly related spotted gar orthologous elements. Three non-conserved elements also play a likely role in wnt1 regulation. The identified enhancers display dynamic modes of chromatin accessibility, suggesting controlled deployment during embryogenesis. Our results suggest that the control of wnt1 and wnt10b expression is under complex regulation involving the interaction of multiple enhancers.

The adoption of cell-based therapies into the clinic will require tremendous large-scale expansio... more The adoption of cell-based therapies into the clinic will require tremendous large-scale expansion to satisfy future demand, and bioreactor-microcarrier cultures are best suited to meet this challenge. The use of spherical microcarriers, however, precludes in-process visualization and monitoring of cell number, morphology, and culture health. The development of novel expansion methods also motivates the advancement of analytical methods used to characterize these microcarrier cultures. A robust optical imaging and image-analysis assay to non-destructively quantify cell number and cell volume was developed. This method preserves 3D cell morphology and does not require membrane lysing, cellular detachment, or exogenous labeling. Complex cellular networks formed in microcarrier aggregates were imaged and analyzed in toto. Direct cell enumeration of large aggregates was performed in toto for the first time. This assay was successfully applied to monitor cellular growth of mesenchymal st...

Experimental Biology and Medicine, 2020

Mutations in ACTA2, encoding smooth muscle α-actin, are a frequent cause of heritable thoracic ao... more Mutations in ACTA2, encoding smooth muscle α-actin, are a frequent cause of heritable thoracic aortic aneurysm and dissections. These mutations are associated with impaired vascular smooth muscle cell function, which leads to decreased ability of the cell to sense matrix-mediated mechanical stimuli. This study investigates how loss of smooth muscle α-actin affects cytoskeletal tension development and cell adhesion using smooth muscle cells explanted from aorta of mice lacking smooth muscle α-actin. We tested the hypothesis that reduced vascular smooth muscle contractility due to a loss of smooth muscle α-actin decreases cellular mechanosensing by dysregulating cell adhesion to the matrix. Assessment of functional mechanical properties of the aorta by stress relaxation measurements in thoracic aortic rings suggested two functional regimes for Acta2−/−mice. Lower stress relaxation was recorded in aortic rings from Acta2−/−mice at tensions below 10 mN compared with wild type, likely dr...

International Journal of Molecular Sciences, 2019

Watermelon fruit contains a high percentage of amino acid citrulline (Cit) and arginine (Arg). Ci... more Watermelon fruit contains a high percentage of amino acid citrulline (Cit) and arginine (Arg). Cit and Arg accumulation in watermelon fruit are most likely mediated by both de novo synthesis from other amino acids within fruits and direct import from source tissues (leaves) through the phloem. The amino acid transporters involved in the import of Cit, Arg, and their precursors into developing fruits of watermelon have not been reported. In this study, we have compiled the list of putative amino acid transporters in watermelon and characterized transporters that are expressed in the early stage of fruit development. Using the yeast complementation study, we characterized ClAAP3 (Cla023187) and ClAAP6 (Cla023090) as functional amino acid transporters belonging to the family of amino acid permease (AAP) genes. The yeast growth and uptake assays of radiolabeled amino acid suggested that ClAAP3 and ClAAP6 can transport a broad spectrum of amino acids. Expression of translational fusion p...

Molecular biology of the cell, Jan 27, 2017

Although it is known that Nck regulates cell adhesion and migration by bridging tyrosine phosphor... more Although it is known that Nck regulates cell adhesion and migration by bridging tyrosine phosphorylation with cytoskeletal remodeling, the role of Nck in tumorigenesis and metastasis has remained undetermined. Here, we report that Nck is required for the growth and vascularization of primary tumors and lung metastases in a breast cancer xenograft model as well as extravasation following injection of carcinoma cells into the tail vein. We provide evidence that Nck directs the polarization of cell-matrix interactions for efficient migration in three-dimensional microenvironments. We show that Nck advances breast carcinoma cell invasion by regulating actin dynamics at invadopodia and enhancing focalized extracellular matrix proteolysis by directing the delivery and accumulation of MMP14 at the cell surface. We find that Nck-dependent cytoskeletal changes are mechanistically linked to enhanced RhoA but restricted spatiotemporal activation of Cdc42. Using a combination of protein silenci...

Journal of biomedical optics, 2014

During embryogenesis, presumptive brain compartments are patterned by dynamic networks of gene ex... more During embryogenesis, presumptive brain compartments are patterned by dynamic networks of gene expression. The spatiotemporal dynamics of these networks, however, have not been characterized with sufficient resolution for us to understand the regulatory logic resulting in morphogenetic cellular behaviors that give the brain its shape. We have developed a new, integrated approach using ultrashort pulse microscopy [a high-resolution, two-photon fluorescence (2PF)-optical coherence microscopy (OCM) platform using 10-fs pulses] and image registration to study brain patterning and morphogenesis in zebrafish embryos. As a demonstration, we used time-lapse 2PF to capture midbrain-hindbrain boundary morphogenesis and a wnt1 lineage map from embryos during brain segmentation. We then performed in situ hybridization to deposit NBT/BCIP, where wnt1 remained actively expressed, and reimaged the embryos with combined 2PF-OCM. When we merged these datasets using morphological landmark registratio...

Optical Engineering, 2013

Lymphatic Research and Biology, 2012

Journal of Biomedical Optics, 2014

Journal of Biomedical Optics, 2013

The mechanisms that enable viruses to harness cellular machinery for their own survival are prima... more The mechanisms that enable viruses to harness cellular machinery for their own survival are primarily studied in cell lines cultured in two-dimensional (2-D) environments. However, there are increasing reports of biological differences between cells cultured in 2-D versus three-dimensional (3-D) environments. Here we report differences in host-virus interactions based on differences in culture environment. Using ultrashort pulse microscopy (UPM), a form of two-photon microscopy that utilizes sub-10-fs pulses to efficiently excite fluorophores, we have shown that de novo development of extra-chromosomal virus replication compartments (VRCs) upon murine cytomegalovirus (mCMV) infection is markedly enhanced when host cells are cultured in 3-D collagen gels versus 2-D monolayers. In addition, time-lapse imaging revealed that mCMV-induced VRCs have the capacity to grow by coalescence. This work supports the future potential of 3-D culture as a useful bridge between traditional monolayer cultures and animal models to study host-virus interactions in a more physiologically relevant environment for the development of effective anti-viral therapeutics. These advances will require broader adoption of modalities, such as UPM, to image deep within scattering tissues.

Frontiers in Neuroanatomy, 2017

Frontiers in physiology, 2018

Aging induces a progressive decline in vasoconstrictor responses in central and peripheral arteri... more Aging induces a progressive decline in vasoconstrictor responses in central and peripheral arteries. This study investigated the hypothesis that vascular smooth muscle (VSM) contractile function declines with age in soleus muscle feed arteries (SFA). Contractile function of cannulated SFA isolated from young (4 months) and old (24 months) Fischer 344 rats was assessed by measuring constrictor responses of denuded (endothelium removed) SFA to norepinephrine (NE), phenylephrine (PE), and angiotensin II (Ang II). In addition, we investigated the role of RhoA signaling in modulation of VSM contractile function. Structural and functional characteristics of VSM cells were evaluated by fluorescence imaging and atomic force microscopy (AFM). Results indicated that constrictor responses to PE and Ang II were significantly impaired in old SFA, whereas constrictor responses to NE were preserved. In the presence of a Rho-kinase inhibitor (Y27632), constrictor responses to NE, Ang II, and PE wer...