HoxB genes regulate neuronal delamination in the trunk neural tube by controlling the expression of Lzts1

Abstract

Differential Hox gene expression is central for specification of axial neuronal diversity in the spinal cord. Here, we uncover an additional function of Hox proteins in the developing spinal cord, restricted to B cluster Hox genes. We found that members of the HoxB cluster are expressed in the trunk neural tube of chicken embryo earlier than Hox from the other clusters, with poor antero-posterior axial specificity and with overlapping expression in the intermediate zone (IZ). Gain-of-function experiments of HoxB4, HoxB8 and HoxB9, respectively representative of anterior, central, and posterior HoxB genes, resulted in ectopic progenitor cells in the mantle zone. The search for HoxB8 downstream targets in the early neural tube identified the Leucine Zipper Tumor Suppressor 1 gene (Lzts1), whose expression is also activated by HoxB4 and HoxB9. Gain and loss of function experiments showed that Lzts1, expressed endogenously in the IZ, controls neuronal delamination. These data collectively indicate that HoxB genes have a generic function in the developing spinal cord, controlling the expression of Lzts1 and neuronal delamination.

Sustained experimental activation of FGF8/ERK in the developing chicken spinal cord reproducibly models early events in ERK-mediated tumorigenesis

“…The whole mount in situ hybridization protocol is as described (Wilmerding et al, 2021). Embryos were fixed 2 hours at RT in 4% buffered formaldehyde in PBS.…”

“…Parts of the neural tube expressing GFP were microdissected one day after electroporation and dissociated with trypsin/EDTA 0.25%. A highly enriched population of GFP-expressing cells was isolated by FACS with the use of a dead cell exclusion (DCE)/discrimination dye (DAPI) to eliminate dying cells (Wilmerding et al, 2021). RNA was extracted (RNeasy Mini Kit), reverse transcribed with random primers and cDNA was amplified using a linear amplification system used for building sequencing libraries (done at the GATC).…”

HOXB8 Counteracts MAPK/ERK Oncogenic Signaling in a Chicken Embryo Model of Neoplasia

“…We have recently analyzed the function and identified target genes of HOXB8 in the developing spinal cord of chicken embryo [12]. In particular, we have demonstrated that the gain of function of HOXB8 in the trunk neural tube of a 2-day-old chicken embryo performed by electroporation of the pCIG-HOXB8 expressing vector (co-expressing nuclear GFP) increases cell death and the amount of the transcript of the gene coding for the tumor suppressor LZTS1 [12]. To test the effect of HOXB8 on ERK oncogenic activity in an in vivo context, we took advantage of a model of ERK-induced neoplasia we developed in the chicken embryo neural tube [27].…”

“…Similarly, HOXB8 acts as an oncogene in several cancers including colorectal cancer [10], but is a favorable prognostic marker in renal cancer (Human Protein Atlas) [11]. HOXB8 has also been recently shown to positively control the transcriptional expression of the tumor suppressor LZTS1 [12]. A better understanding of the molecular mechanisms that promote either the oncogenic or tumor suppressor activity of the HOX proteins remains unsolved.…”

“…To gain insights in the molecular mechanisms by which the HOX proteins control ERK oncogenic pathways, we developed a chicken embryo neoplasia model obtained by ERK1/2 overactivation in the trunk neural tube [27] and investigated the effects of HOX expression in this model. We started the analysis with HOXB8, for which we have recently investigated the function during the development of the spinal cord and identified transcriptional target genes in this structure [12]. We found that HOXB8 largely counteracts ERK oncogenic activity in this neoplasia model.…”