Biology subjects

Nechiporuk, A. V.

Publications and source records attributed to Nechiporuk, A. V..

3 recordsLinked to original sources

bioRxiv2023

RhoA GEF Mcf2lb regulates rosette integrity during collective cell migration

During development, multicellular rosettes serve as important cellular intermediates in the formation of diverse organ systems. Multicellular rosettes are transient epithelial structures that are defined by the apical constriction of cells towards the rosette center. Due to the important role these structures play during development, understanding the molecular mechanisms by which rosettes are formed and maintained is of high interest. Utilizing the zebrafish posterior lateral line primordium (pLLP) as a model system, we identify the RhoA GEF Mcf2lb as a regulator of rosette integrity. The pLLP is a group of [~]150 cells that migrates along the zebrafish trunk and is organized into epithelial rosettes; these are deposited along the trunk and will differentiate into sensory organs called neuromasts (NMs). Using single-cell RNA sequencing and whole-mount in situ hybridization, we showed that mcf2lb is expressed in the pLLP during migration. Given the known role of RhoA in rosette formation, we asked whether Mcf2lb plays a role in regulating apical constriction of cells within rosettes. Live imaging and subsequent 3D analysis of mcf2lb mutant pLLP cells showed disrupted apical constriction and subsequent rosette organization. This in turn resulted in a unique posterior Lateral Line phenotype: an excess number of deposited NMs along the trunk of the zebrafish. Cell polarity markers ZO-1 and Par-3 were apically localized, indicating that pLLP cells are normally polarized. In contrast, signaling components that mediate apical constriction downstream of RhoA, Rock-2a and non-muscle Myosin II were diminished apically. Altogether our results suggest a model whereby Mcf2lb activates RhoA, which in turn activates downstream signaling machinery to induce and maintain apical constriction in cells incorporated into rosettes.

developmental biology↗

bioRxiv2021

c-Kit mediates cutaneous sensory axon innervation and multi-kinase inhibitor-induced neurotoxicity

Peripheral somatosensory neurons innervate the skin and sense the environment. Loss of skin innervation, often caused by the "dying back" of distal somatosensory axons, is a common side effect of drug-induced peripheral neuropathies (DIPNs) and results in pain and sensory dysfunction. Targeted cancer therapies frequently employ multi-kinase inhibitor (MKI) drugs that each block multiple receptor tyrosine kinases. Many MKIs produce DIPNs but the molecular targets and cellular mechanisms underlying these are unknown. We performed live-imaging of cutaneous somatosensory axons in larval zebrafish during treatment with several MKIs known to induce DIPNs, and observed axonal retraction consistent with a dying back pathology. These results were replicated in mouse somatosensory neurons. Genetic knockout of potential MKI targets identified c-Kit receptor as a regulator of sensory axon innervation and a major target of these MKIs mediating loss of axonal density. In both fish and mammals, Kit receptor is expressed in cutaneous somatosensory neurons and its ligand, Kitlg, is expressed in the skin. Mosaic misexpression of Kitlg in the skin induced dramatic increases in local sensory axon density, suggesting an important role for Kit signaling in cutaneous axon growth and maintenance. Immunostaining and structure-function analysis revealed Src, a downstream Kit target, mediates Kits role in cutaneous axon innervation and MKI neurotoxicity. Our data shows that the Kit-Src signaling pathway has a major role in cutaneous sensory axon innervation and is a potential therapeutic target to address DIPNs caused by MKIs and other compounds.

neuroscience↗

bioRxiv2018

A specific Ret receptor isoform is required for pioneer axon outgrowth and growth cone dynamics

In many cases, axon growth and guidance are driven by pioneer axons, the first axons to grow in a particular region. Despite their dynamic pathfinding capabilities and developmental importance, there are very few pioneer neuron specific markers and thus their in vivo identification and functional interrogation have been difficult. We found that a Ret receptor isoform, Ret51, is highly enriched in peripheral sensory pioneer neurons and is required for pioneer axon outgrowth. Ret null mutant pioneer neurons differentiate normally; however, they displayed defects in growth cone morphology and formation of filopodia before pioneer axon extension prematurely halts. We also demonstrate loss-of-function of a retrograde cargo adaptor, JNK-interacting protein 3 (Jip3), phenocopied many of these axonal defects. We further found that loss of Jip3 led to accumulation of activated Ret receptor in pioneer growth cones, indicating a failure in the clearance of activated Ret from growth cones. Using an axon sever approach as well as in vivo analysis of axonal transport, we showed Jip3 specifically mediates retrograde, but not anterograde, transport of activated Ret51. Finally, live imaging revealed that Jip3 and Ret51 were retrogradely co-transported in pioneer axons, suggesting Jip3 functions as an adapter for retrograde transport of Ret51. Taken together, these results identify Ret51 as a molecular marker of pioneer neurons and elucidate an important isoform-specific role for Ret51 in axon growth and growth cone dynamics during development.

developmental biology↗