Biology subjects

Reyes-Nava, N. G.

Publications and source records attributed to Reyes-Nava, N. G..

6 recordsLinked to original sources

bioRxiv2026

Odad1 is Required for Deployment of Axonemal Outer Arm Dyneins and Embryonic Left-Right Patterning

Motile cilia generate fluid flow required for development and tissue homeostasis. Defects in ciliary motility cause Primary Ciliary Dyskinesia (PCD), presenting with situs inversus, infertility, chronic respiratory disease, and hydrocephalus. Ciliary beating requires outer dynein arms (ODAs), anchored to microtubules by the ODA docking complex (ODA-DC). ODAD1 is an ODA-DC component and a known PCD protein. ODAD1 alleles are often associated with human laterality defects, however Odad1 has not been reported to localize to 9+0 axonemes. Here, we show that GFP-Odad1 localizes to nodal cilia (9+0) in the gastrocoel roof plate (GRP), the frog's equivalent to the mammalian node where left-right asymmetry is first established. We found that Odad1 knockdown (KD) disrupts left-right patterning in Xenopus, with loss of pitx2 expression and abnormal gut looping reflecting defective 9+0 nodal cilia at the GRP. In 9+2 multiciliated cells (MCC) axonemes, Odad1 loss impaired deployment of ODA motors and ODA-DC subunits, disrupting ciliary motility. Two patient-derived truncating ODAD1 alleles were similarly unstable, failing to rescue ODA docking in MCCs or left-right patterning at the GRP. Together, these results show ODAD1 is required for ODA-DC assembly in both 9+0 and 9+2 cilia, linking its loss to ciliary motility and left-right patterning defects in ODAD1-associated ciliopathy.

developmental biology↗

bioRxiv2025

Physical and functional interaction of Lrrc56 and Odad3 controls deployment of axonemal dyneins in vertebrate multiciliated cells

Primary ciliary dyskinesia is a genetically heterogeneous motile ciliopathy characterized chronic respiratory disease, laterality defects, hydrocephalus, and infertility, caused by impaired function of motile cilia. LRRC56 has recently emerged as a novel PCD candidate gene, but its role in vertebrate cilia remains poorly understood. Here, we used Xenopus laevis multiciliated cells, targeted knockdown, and in vivo imaging to investigate Lrrc56 function. We show that loss of lrrc56 causes specific depletion of outer dynein arms (ODAs) from the distal axoneme. In vivo affinity purification mass spectrometry revealed that Lrrc56 binds the ODA docking complex components, including Odad3. Consistently, Lrrc56 knockdown also led to distal loss of Odad3. Moreover, we show that disease-associated variants in LRRC56 and ODAD3 disrupted their localization and interaction, pointing to a shared functional pathway. Our work demonstrates that Lrrc56 is a critical regulator of distal ODAs and ODA docking complex deployment and provides new mechanistic insight into how LRRC56 mutations contribute to PCD.

cell biology↗

bioRxiv2023

Mutation of gabra1 is associated with hypermotility and abnormal expression of proteins critical for ion homeostasis and synaptic vesicle transport.

Mutation of the GABRA1 gene is associated with neurodevelopmental defects and epilepsy. GABRA1 encodes for the 1 subunit of the gamma-aminobutyric acid type A receptor (GABAAR), which regulates the fast inhibitory impulses of the nervous system. Multiple model systems have previously been developed to understand the function of GABRA1 during development, but these models have produced complex and at times incongruent data. Thus, additional model systems are required to validate and substantiate previously published results. We investigated the behavioral swim patterns associated with a nonsense mutation of the zebrafish gabra1 (sa43718 allele) gene. The sa43718 allele causes a decrease in gabra1 mRNA expression, which is associated with light induced hypermotility, one phenotype associated with seizure like behavior in zebrafish. Mutation of gabra1 was accompanied by decreased mRNA expression of gabra2, gabra3, and gabra5, indicating a reduction in the expression of additional alpha sub-units of the GABAAR. Although multiple sub-units were decreased in total expression, larvae continued to respond to pentylenetetrazole (PTZ) indicating that a residual GABAAR exists in the sa43718 allele. Proteomics analysis demonstrated that nonsense mutation of gabra1 is associated with abnormal expression of proteins that regulate proton transport, ion homeostasis, vesicle transport, and mitochondrial protein complexes. These data support previous studies performed in a zebrafish nonsense allele created by CRISPR/Cas9 and validate that loss of function mutations in the gabra1 gene result in seizure like phenotypes with abnormal function of inhibitory synapses.

developmental biology↗

bioRxiv2023

Abnormal chondrocyte intercalation in a zebrafish model of cblC syndrome restored by an MMACHC cobalamin binding mutant.

Variants in the MMACHC gene cause combined methylmalonic acidemia and homocystinuria cblC type, the most common inborn error of intracellular cobalamin (vitamin B12) metabolism. cblC is associated with neurodevelopmental, hematological, ocular, and biochemical abnormalities. In a subset of patients, mild craniofacial dysmorphia has also been described. Mouse models of Mmachc deletion are embryonic lethal but cause severe craniofacial phenotypes such as facial clefts. MMACHC encodes an enzyme required for cobalamin processing and variants in this gene result in the accumulation of two metabolites: methylmalonic acid (MMA) and homocysteine (HC). Interestingly, other inborn errors of cobalamin metabolism, such as cblX syndrome, are associated with mild facial phenotypes. However, the presence and severity of MMA and HC accumulation in cblX syndrome is not consistent with the presence or absence of facial phenotypes. Thus, the mechanisms by which mutation of MMACHC cause craniofacial defects have not been completely elucidated. Here we have characterized the craniofacial phenotypes in a zebrafish model of cblC (hg13) and performed restoration experiments with either wildtype or a cobalamin binding deficient MMACHC protein. Homozygous mutants did not display gross morphological defects in facial development, but did have abnormal chondrocyte intercalation, which was fully penetrant. Abnormal chondrocyte intercalation was not associated with defects in the expression/localization of neural crest specific markers, sox10 or barx1. Most importantly, chondrocyte organization was fully restored by wildtype MMACHC and a cobalamin binding deficient variant of MMACHC protein. Collectively, these data suggest that mutation of MMACHC causes mild to moderate craniofacial phenotypes that are independent of cobalamin binding.

developmental biology↗

bioRxiv2020

Activation of WNT signaling restores the facial deficits in a zebrafish with defects in cholesterol metabolism.

BackgroundInborn errors of cholesterol metabolism occur as a result of mutations in the cholesterol synthesis pathway (CSP). Although mutations in the CSP cause a multiple congenital anomaly syndrome, craniofacial abnormalities are a hallmark phenotype associated with these disorders. Previous studies have established that mutation of the zebrafish hmgcs1 gene (Vu57 allele), which encodes the first enzyme in the CSP, causes defects in craniofacial development and abnormal neural crest cell (NCC) differentiation. However, the molecular mechanisms by which the products of the CSP disrupt NCC differentiation are not completely known. Cholesterol is known to regulate the activity of WNT signaling, an established regulator of NCC differentiation. We hypothesized that defects in cholesterol synthesis reduce WNT signaling, consequently resulting in abnormal craniofacial development. MethodsTo test our hypothesis we performed a combination of pharmaceutical inhibition, gene expression assays, and targeted rescue experiments to understand the function of CSP and WNT signaling during craniofacial development. ResultsWe demonstrate reduced expression of axin2, a WNT downstream target gene in homozygous carriers of the Vu57 allele and in larvae treated with Ro-48-8071, which inhibits the synthesis of cholesterol. Moreover, activation of WNT signaling via treatment with a WNT agonist completely restored the craniofacial defects present in the Vu57 allele. ConclusionsCollectively, these data suggest interplay between the CSP and WNT signaling during craniofacial development.

developmental biology↗

bioRxiv2020

Abnormal expression of GABAA receptor sub-units and hypomotility upon loss of gabra1 in zebrafish.

We used whole exome sequencing (WES) to determine the genetic etiology of a patient with a multi-system disorder characterized by a seizure phenotype. WES identified a heterozygous de novo missense mutation in the GABRA1 gene (c.875C>T). GABRA1 encodes the alpha subunit of the Gamma-Aminobutyric Acid receptor A (GABAAR). The GABAAR is a ligand gated ion channel that mediates the fast inhibitory signals of the nervous system and mutations in the sub-units that compose the GABAAR have been previously associated with human disease. To understand the mechanisms by which GABRA1 regulates brain development, we developed a zebrafish model of gabra1 deficiency. gabra1 expression is restricted to the nervous system and behavioral analysis of morpholino injected larvae suggests that the knockdown of gabra1 results in hypoactivity and defects in the expression of other sub-units of the GABAAR. Expression the human GABRA1 protein in morphants partially restored the hypomotility phenotype. In contrast, the expression of the c.875C>T variant did not restore these behavioral deficits. Collectively, these results represent a functional approach to understand the mechanisms by which loss of function alleles cause disease.

developmental biology↗