Biology subjects

Gruner, H. N.

Publications and source records attributed to Gruner, H. N..

4 recordsLinked to original sources

bioRxiv2025

Neural signaling contributes to heart formation and growth in the invertebrate chordate, Ciona robusta.

Neurons contribute to the complex interplay of signals that mediate heart development and homeostasis. Although a limited set of studies suggest that neuronal peptides impact vertebrate heart growth, the specific contributions of these peptides to cardiomyocyte progenitor differentiation or proliferation have not been elucidated. Here we show that the neuropeptide tachykinin along with canonical Wnt signaling regulate cardiomyocyte progenitor proliferation in the chordate model Ciona robusta. In C. robusta, the heart continues to grow throughout adulthood and classic histological studies indicate that a line of undifferentiated cells may serve as a reserve progenitor lineage. We found that this line of cardiomyocyte progenitors consists of distinct distal and midline populations. Distal progenitors divide asymmetrically to produce distal and midline daughters. Midline progenitors divide asymmetrically to produce myocardial precursors. Through single cell RNA sequencing (scRNA-seq) of adult C. robusta hearts, we delineated the cardiomyocyte progenitor expression profile. Based on this data we investigated the role of Wnt signaling in cardiomyocyte progenitor proliferation and found that canonical Wnt signaling is required to suppress excessive progenitor proliferation. The scRNA-seq data also identified a number of presumptive cardiac neural-like cells. Strikingly, we found that a subset of these neuronal cells appears to innervate the distal cardiomyocyte progenitors. Based on the expression of the tachykinin receptor in these neuronal cells, we blocked tachykinin signaling using pharmacological inhibitors and found that this drove reduced proliferation in the distal progenitor pool. Through targeted CRISPR-Cas9 knockdown we then demonstrated that both extrinsic tachykinin and intrinsic, cardiac tachykinin receptors are required for formation of the myocardial heart tube. This work provides valuable insights into how organisms may deploy neural signals to regulate organ growth in response to environmental or homeostatic inputs.

developmental biology↗

bioRxiv2019

Precise removal of Calm1 long 3′ UTR isoform by CRISPR-Cas9 genome editing impairs dorsal root ganglion development in mice.

Most mammalian genes are subject to Alternative cleavage and PolyAdenylation (APA), often resulting in alternative length 3' UTR isoforms. Thousands of extended or long 3' UTR variants are preferentially expressed in neuron-enriched tissues of metazoans. However, the in vivo functions of these long 3' UTR isoforms are largely unknown. Calmodulin 1 (Calm1) is a key integrator of calcium signaling that is required for correct neural development. Calm1 generates short (Calm1-S) and long 3' UTR (Calm1-L) mRNA isoforms via APA. We found Calm1-S to be broadly expressed across mouse tissues, whereas Calm1-L expression was largely restricted to neural tissues, including the dorsal root ganglion (DRG). Using CRISPR-Cas9 genome editing, a series of mouse deletion lines were generated that successfully eliminated expression of Calm1-L while maintaining expression of Calm1-S. One of these lines, Calm1{Delta}3' UTR, carried a 163 bp deletion surrounding the distal polyA site. Examination of Calm1{Delta}3' UTR embryos revealed disrupted development of the DRG. In Calm1{Delta}3' UTR DRG explant cultures undergoing axon outgrowth, we observed a dramatic increase in axon fasciculation. These results demonstrate a physiological role for Calm1-L in DRG development, and more generally, establish a genome-editing strategy to study in vivo functions of long 3' UTR isoforms. Author SummaryMore than half of all human genes generate alternative mRNA isoforms which differ in the length of their 3 Untranslated regions (3 UTRs). Through a process called Alternative Cleavage and Polyadenylation thousands of broadly expressed genes preferentially express long 3 UTR variants in brain tissues whereas their short 3 UTR counterparts are more broadly expressed. A challenge to study the functions of these transcripts has been to generate loss of function mutant animals that lack a long 3 UTR isoform but maintain expression of the corresponding short 3 UTR isoform. Here, we used the precise, rapid, and efficient approach of CRISPR genome-editing to generate long 3 UTR mutant mice. These mice, which do not express the long 3 UTR of the Calmodulin 1 (Calm1) gene, exhibit impairment in the development of sensory neurons, including increased fasciculation of axons and aberrant cell body migration. This finding is important because it provides conclusive genetic evidence for a neural function of a long 3 UTR isoform in an animal. The CRISPR genome-editing approach used here can be applied to the study of neuron-enriched long 3 UTR isoforms, which number in the thousands and have largely unexplored functions.

genetics↗

bioRxiv2018

Robo1 and 2 repellent receptors cooperate to guide facial neuron cell migration and axon projections in the embryonic mouse hindbrain

BackgroundThe facial nerve is necessary for our ability to eat, speak, and make facial expressions. Both the axons and cell bodies of the facial nerve undergo a complex embryonic migration pattern involving migration of the cell bodies caudally and tangentially through rhombomeres, and simultaneously the axons projecting to exit the hindbrain to form the facial nerve.\n\nResultsOur goal in this study was to test the functions of the chemorepulsive receptors Robo1 and Robo2 in facial neuron and axon migration by analyzing genetically marked motor neurons in double mutant mouse embryos through the migration time course, E10.0-E13.5. In Robo1/2 double mutants, axon and cell body migration errors were more severe than in single mutants. Most axons did not make it to their motor exit point, and instead projected into and longitudinally within the floor plate. Surprisingly, some facial neurons had bifurcated axons that either exited or projected into the floor plate. At the same time, a subset of mutant facial cell bodies failed to migrate caudally, and instead shifted into the floor plate.\n\nConclusionsRobo1 and Robo2 have redundant functions to guide multiple aspects of the complex cell migration of the facial nucleus, as well as regulating axon trajectories and suppressing formation of ectopic axons.\n\nBullet pointsO_LIRobo1 and 2 repellent receptors were tested for functions in facial neuron cell body and axon migration in the developing E10.0-E13.5 hindbrain of double mutant mouse embryos.\nC_LIO_LIIn Robo1;Robo2 double mutant embryos, facial motor neuron cell bodies and axons abnormally grew into the hindbrain floor plate, with stronger effects than in single Robo mutants.\nC_LIO_LIMany facial nerve axons abnormally bifurcated, failed to project to their proper exit point, and instead projected longitudinally within the floor plate.\nC_LIO_LIFacial neuron cell body tangential migration was disrupted.\nC_LIO_LIRobo receptor-mediated repulsion is required for proper migration of both facial axon projections and cell body migrations.\nC_LI\n\nFunding was provided to G.S.M. by NIH R01 NS054740, R21 NS077169, and R01 EY025205. Core facilities at the University of Nevada Reno campus are supported by NIH COBREs GM103650 and GM103554, and Nevada INBRE GM103440.

neuroscience↗

bioRxiv2017

Global accumulation of circRNAs during aging in Caenorhabditis elegans

Circular RNAs (CircRNAs) are a newly appreciated class of RNAs that lack free 5{acute} and 3{acute} ends, are expressed by the thousands in diverse forms of life, and are mostly of enigmatic function. Ostensibly due to their resistance to exonucleases, circRNAs are known to be exceptionally stable. Here, we examined the global profile of circRNAs in C. elegans during aging by performing ribo-depleted total RNA-seq from the fourth larval stage (L4) through 10-day old adults. Using stringent bioinformatic criteria and experimental validation, we annotated 1,166 circRNAs, including 575 newly discovered circRNAs. These circRNAs were derived from 797 genes with diverse functions, including genes involved in the determination of lifespan. A massive accumulation of circRNAs during aging was uncovered. Many hundreds of circRNAs were significantly increased among the aging time-points and increases of select circRNAs by over 40-fold during aging were quantified by qRT-PCR. The age-accumulation of circRNAs was not accompanied by increased expression of linear RNAs from the same host genes. We attribute the global scale of circRNA age-accumulation to the high composition of postmitotic cells in adult C. elegans, coupled with the high resistance of circRNAs to decay. These findings suggest that the exceptional stability of circRNAs might explain age-accumulation trends observed from neural tissues of other organisms, which also have a high composition of post-mitotic cells. Given the suitability of C. elegans for aging research, it is now poised as an excellent model system to determine if there are functional consequences of circRNA accumulation during aging.

genomics↗