Early expression of Tubulin Beta-III in avian cranial neural crest cells

Abstract

Neural crest cells are a transient stem-like cell population that forms in the dorsal neural tube of vertebrate embryos and then migrates to various locations to differentiate into diverse derivatives such as craniofacial bone, cartilage, and the enteric and peripheral nervous systems. The current dogma of neural crest cell development suggests that there is a specific hierarchical gene regulatory network (GRN) that controls the induction, specification, and differentiation of these cells at specific developmental times. Our lab has identified that a marker of differentiated neurons, Tubulin Beta-III (TUBB3), is expressed in premigratory neural crest cells. TUBB3 has previously been identified as a major constituent of microtubules and is required for the proper guidance and maintenance of axons during development. Using the model organism, Callus gallus, we have characterized the spatiotemporal localization of TUBB3 in early stages of development. Here we show TUBB3 is expressed in the developing neural plate, is upregulated in the pre-migratory cranial neural crest prior to cell delamination and migration, and it is maintained or upregulated in neurons in later developmental stages. We believe that TUBB3 likely has a role in early neural crest formation and migration separate from its role in neurogenesis.

RNA-binding protein Elavl1/HuR is required for maintenance of cranial neural crest specification

“…For cross-sections, embryos were fixed at room temperature for 1 hr, then washed, embedded, and cryosectioned as described ( Hutchins and Bronner, 2018 ; Hutchins and Bronner, 2019 ) prior to immunohistochemistry. Washes, blocking (10% donkey serum), and antibody incubations were performed in TBSTx (0.5 M Tris–HCl/1.5 M NaCl/10 mM CaCl 2 /0.5% Triton X-100/0.001% Thimerosal) as described ( Chacon and Rogers, 2019 ; Manohar et al, 2020 ). Primary antibodies are listed in the Key Resources Table.…”

βIII-Tubulin Gene Regulation in Health and Disease

“…This notion has been strengthened by the identification of mutations in TUBB3 , the gene that encodes for βIII-tubulin, resulting in nervous system disorders such as Congenital Fibrosis of the Extraocular Muscles type 3 (CFEOM3), which combines the weakening of the extraocular muscles with intellectual disability, as well as axonal abnormalities and disorganisation of cortical neurons ( Poirier et al, 2010 ; Tischfield et al, 2010 ). Evidence also suggests βIII-tubulin has roles outside of neurogenesis, such as the formation of neural crest cell formation ( Haendel et al, 1996 ; Chacon and Rogers 2019 ) and recently, in the mineralisation stages of tooth development ( Oshima and Yawaka 2020 ). As such, despite the original neuronal findings, βIII-tubulin expression is increasingly observed outside of neuronal tissue, with reports of adult stem cells expressing βIII-tubulin, such as melanocytes ( Locher et al, 2013 ) and spermatogenic cells ( Person et al, 2017 ).…”

“…While the factors involving Tubb3 expression described in the preceding section are associated with neuronal differentiation of the CNS, it is uncertain whether these shared by differentiating neurons of the PNS. During fetal development, however, βIII-tubulin has been shown to be expressed by neural crest cell progenitors (Pax7 positive cells), and by pre-migratory neural crest cells (Sox9 and Slug positive cells) prior to neurogenesis in the CNS ( Chacon and Rogers 2019 ). Neural crest cells are considered to be multipotent progenitor cells able to give rise to various cells including neurons and melanocytes, and form the majority of the PNS ( Acloque et al, 2008 ).…”

“…Sox11 had the greatest impact on βIII-tubulin expression in neurogenesis ( Bergsland et al, 2006 ; Hoser et al, 2008 ), and is required for binding to NeuroG1 in order to promote Tubb3 in early-born neurons, a process that can be inhibited through Bm2 ( Chen et al, 2015 ). Since these three genes have been linked to neural crest formation ( Uy et al, 2015 ), it is therefore plausible to think that the expression of Tubb3 observed by the neural crest progenitors ( Chacon and Rogers 2019 ) could be driven by members of the SoxC family.…”

“…Slug is co-expressed with βIII-tubulin and Sox9 in pre-migratory avian neural crest cells ( Chacon and Rogers 2019 ). Additionally, SLUG has been shown to directly interact with SOX9 to promote the formation of cancer stem-like cells in lung cancer ( Luanpitpong et al, 2016 ), and there is the recent speculation that TUBB3 may be playing a role in the maintenance of cancer stem like cells ( Namekawa et al, 2020 ).…”