The enteric nervous system of C. elegans is specified by the Sine Oculis-like homeobox gene ceh-34

Overarching themes in the terminal differentiation of the enteric nervous system, an autonomously acting unit of animal nervous systems, have so far eluded discovery. We describe here the overall regulatory logic of enteric nervous system differentiation of the nematode C. elegans that resides within the foregut (pharynx) of the worm. A Caenorhabditis elegans homolog of the Drosophila Sine Oculis homeobox gene, ceh-34, is expressed in all 14 classes of interconnected pharyngeal neurons from their birth throughout their life time, but in no other neuron type of the entire animal. Constitutive and temporally controlled ceh-34 removal shows that ceh-34 is required to initiate and maintain the neuron type-specific terminal differentiation program of all pharyngeal neuron classes, including their circuit assembly, without affecting panneuronal features. Through additional genetic loss of function analysis, we show that within each pharyngeal neuron class, ceh-34 cooperates with different homeodomain transcription factors to individuate distinct pharyngeal neuron classes. Our analysis underscores the critical role of homeobox genes in neuronal identity specification and links them to the control of neuronal circuit assembly of the enteric nervous system. Together with the pharyngeal nervous system simplicity as well as its specification by a Sine Oculis homolog, our findings invite speculations about the early evolution of nervous systems.

The Role of Lymphatic Marker Prox-1 in Relation to Brain Tumours

The homeobox gene, Prox-1 is a transcription factor essential for lymphatic development (lymphangiogenesis) during embryogenesis. It also performs different functions in various tissues such as: retina, lens, liver, pancreas and the central nervous system. Intense expression of Prox-1 has been demonstrated in the developing spinal cord and brain. In adulthood its expression continues in the hippocampus and cerebellum. In adult tissues the process of lymphatic vasculature formation is accompanied under certain pathological conditions such as inflammation, tissue repair and tumour growth. Prox-1 expression is typical for lymphatic vessels; thus it belongs to one of the most specific and widely used mammalian lymphatic endothelial marker in the detection of lymphangiogenesis and lymphatic vessel invasion in oncogenesis. It has been shown that Prox-1 is involved in cancer development and progression. It’s tumour suppressive and oncogenic properties are proven in several human cancers, including brain tumours. Among all body cancers the brain tumours represent the most feared tumours with very limited treatment options and a poor diagnosis. The aim of this paper was to show the current knowledge of the gene Prox-1 with an emphasis on brain tumours, especially in gliomas.

Concordant Androgen-Regulated Expression of Divergent Rhox5 Promoters in Sertoli Cells

Concordant transcriptional regulation can generate multiple gene products that collaborate to achieve a common goal. Here we report a case of concordant transcriptional regulation that instead drives a single protein to be produced in the same cell type from divergent promoters. This gene product—the RHOX5 homeobox transcription factor—is translated from 2 different mRNAs with different 5′ untranslated regions (UTRs) transcribed from alternative promoters. Despite the fact that these 2 promoters—the proximal promoter (Pp) and the distal promoter (Pd)—exhibit different patterns of tissue-specific activity, share no obvious sequence identity, and depend on distinct transcription factors for expression, they exhibit a remarkably similar expression pattern in the testes. In particular, both depend on androgen signaling for expression in the testes, where they are specifically expressed in Sertoli cells and have a similar stage-specific expression pattern during the seminiferous epithelial cycle. We report evidence for 3 mechanisms that collaborate to drive concordant Pp/Pd expression. First, both promoters have an intrinsic ability to respond to androgen receptor and androgen. Second, the Pp acts as an enhancer to promote androgen-dependent transcription from the Pd. Third, Pd transcription is positively autoregulated by the RHOX5 protein, which is first produced developmentally from the Pp. Together, our data support a model in which the Rhox5 homeobox gene evolved multiple mechanisms to activate both of its promoters in Sertoli cells to produce Rhox5 in an androgen-dependent manner during different phases of spermatogenesis.

A SIX1/EYA2 Inhibitor Impairs CALM-AF10 and Jurkat Leukemia Cell Proliferation

Background : The CALM-AF10 translocation is found in 5-10% of T-cell acute lymphoblastic leukemias (T-ALL), and a subset of acute myeloid leukemias (AML). CALM-AF10 leukemias are characterized by elevated expression of proleukemic HOXA genes. Since HOXA genes are difficult to target, we hypothesized that identification of non-HOXA CALM-AF10 effector genes could potentially yield novel therapeutic targets. To discover novel CALM-AF10-regulated genes, we took advantage of our prior observation that the nuclear export factor CRM1/XPO1 tethers CALM-AF10 to HOXA genes by interacting with a nuclear export signal within CALM. Using microarrays, we identified a set of genes that showed decreased expression in response to the CRM1 inhibitor Leptomycin B (LMB), similar to Hoxa genes, in murine CALM-AF10 leukemia cells. Then using RNA-sequencing, we discovered a set of genes increased in murine hematopoietic stem cells transduced with CALM-AF10. There were 11 genes that were both decreased in response to LMB and increased in response to CALM-AF10, which included the Hoxa gene cluster, as well as Six1. We demonstrated that CALM-AF10 increases Six1 expression and localizes to the Six1 locus, as it does the Hoxa genes. SIX1, like the Hoxa genes, is a homeobox gene that is associated with embryogenesis and is quiescent post-embryologically. In addition, SIX1 and its cofactor EYA2 are overexpressed in numerous solid tumors, and an inhibitor of the SIX1/EYA2 complex (Compound 8430) has recently been described. While there is evidence of a role for SIX1 in solid tumors, its role in leukemias has not been explored. Objective : Evaluate the effect of a SIX1/EYA2 complex inhibitor on leukemia cell proliferation. Design/Methods : SIX1 gene and protein expression were assessed in CALM-AF10, Jurkat (T-ALL) and NOMO1 (AML) leukemia cell lines via Western Blot and RT-qPCR. CALM-AF10 leukemias were derived from murine models in our lab, Jurkat and NOMO1 cell lines were obtained from ATCC. The effect of compound 8430 - an inhibitor of the Six1/Eya2 interaction - on cell proliferation was evaluated using Cell-Titer-Glo Assays and liquid culture proliferation assays. In addition, we used the the CRM1 Nuclear Export Inhibitor KPT-330 alone and in combination with 8430 in these cell lines. SynergyFinder2 (https://synergyfinder.fimm.fi/) was used to assess synergy of 8430 and KPT-330. δ-score is a calculated value that indicates synergistic drug interaction, with a higher δ-score indicative of a synergistic effect of the drugs. Results : SIX1 gene and protein expression are increased in CALM-AF10 leukemia cell lines and Jurkat T-ALL cells, but not NOMO1 cells. Compound 8430 decreases cell proliferation in CALM-AF10 leukemias and Jurkat leukemia cell lines, however it did not affect the AML line NOMO1. Correspondingly, liquid cultures showed that 8430 alone slowed the proliferation of CALM-AF10 leukemia and the Jurkat cells, but not NOMO1 cells. The addition of KPT-330 to 8430 was synergistic in CALM-AF10 leukemia cells with a KPT-330 dose of 60 nM and multiple dose levels of 8430 (δ-scores 17-19) while in the Jurkat leukemia cells a dose of 30 nM of KPT-330 was synergistic at multiple dose levels of 8430 (δ-score 6-8) (Figure 1). Conclusions : The SIX1 homeobox gene is highly expressed during development, and its expression is silenced post-embryogenesis. Through an initial unbiased screen, we discovered that Six1 may play a role in CALM-AF10 leukemogenesis. We have determined that Six1 expression is upregulated in the presence of CALM-AF10. A role for Six1 in CALM-AF10 leukemogenesis is further supported by the ability of a SIX1/EYA2 inhibitor to slow the proliferation of CALM-AF10 leukemia cells. Importantly, based on our observation that 8430 slows proliferation of Jurkat cells, SIX1 inhibition may be relevant in other leukemias. Finally, our demonstration that 8430 synergizes with KPT-330, a Nuclear Export Inhibitor, suggests the possibility of a novel therapeutic approach for CALM-AF10 and other leukemias. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

A medullary centre for lapping in mice

It has long been known that orofacial movements for feeding can be triggered, coordinated, and often rhythmically organized at the level of the brainstem, without input from higher centers. We uncover two nuclei that can organize the movements for ingesting fluids in mice. These neuronal groups, IRtPhox2b and Peri5Atoh1, are marked by expression of the pan-autonomic homeobox gene Phox2b and are located, respectively, in the intermediate reticular formation of the medulla and around the motor nucleus of the trigeminal nerve. They are premotor to all jaw-opening and tongue muscles. Stimulation of either, in awake animals, opens the jaw, while IRtPhox2b alone also protracts the tongue. Moreover, stationary stimulation of IRtPhox2b entrains a rhythmic alternation of tongue protraction and retraction, synchronized with jaw opening and closing, that mimics lapping. Finally, fiber photometric recordings show that IRtPhox2b is active during volitional lapping. Our study identifies one of the subcortical nuclei underpinning a stereotyped feeding behavior.

Zebrafish Paralogs brd2a and brd2b Are Needed for Proper Circulatory, Excretory and Central Nervous System Formation and Act as Genetic Antagonists during Development

Brd2 belongs to the BET family of epigenetic transcriptional co-regulators that act as adaptor-scaffolds for the assembly of chromatin-modifying complexes and other factors at target gene promoters. Brd2 is a protooncogene and candidate gene for juvenile myoclonic epilepsy in humans, a homeobox gene regulator in Drosophila, and a maternal-zygotic factor and cell death modulator that is necessary for normal development of the vertebrate central nervous system (CNS). As two copies of Brd2 exist in zebrafish, we use antisense morpholino knockdown to probe the role of paralog Brd2b, as a comparative study to Brd2a, the ortholog of human Brd2. A deficiency in either paralog results in excess cell death and dysmorphology of the CNS, whereas only Brd2b deficiency leads to loss of circulation and occlusion of the pronephric duct. Co-knockdown of both paralogs suppresses single morphant defects, while co-injection of morpholinos with paralogous RNA enhances them, suggesting novel genetic interaction with functional antagonism. Brd2 diversification includes paralog-specific RNA variants, a distinct localization of maternal factors, and shared and unique spatiotemporal expression, providing unique insight into the evolution and potential functions of this gene.

Genome-Wide Analysis of the Homeobox Gene Family and Identification of Drought-Responsive Members in Populus trichocarpa

Homeobox (HB) genes play critical roles in the regulation of plant morphogenesis, growth and development. Here, we identified a total of 156 PtrHB genes from the Populus trichocarpa genome. According to the topologies and taxonomy of the phylogenetic tree constructed by Arabidopsis thaliana HB members, all PtrHB proteins were divided into six subgroups, namely HD-ZIP, ZF-HD, HB-PHD, TALE, WOX and HB-OTHERS. Multiple alignments of conserved homeodomains (HDs) revealed the conserved loci of each subgroup, while gene structure analysis showed similar exon–intron gene structures, and motif analysis indicated the similarity of motif number and pattern in the same subgroup. Promoter analysis indicated that the promoters of PtrHB genes contain a series of cis-acting regulatory elements involved in responding to various abiotic stresses, indicating that PtrHBs had potential functions in these processes. Collinearity analysis revealed that there are 96 pairs of 127 PtrHB genes mainly distributing on Chromosomes 1, 2, and 5. We analyzed the spatio-temporal expression patterns of PtrHB genes, and the virus-induced gene silencing (VIGS) of PtrHB3 gene resulted in the compromised tolerance of poplar seedlings to mannitol treatment. The bioinformatics on PtrHB family and preliminary exploration of drought-responsive genes can provide support for further study of the family in woody plants, especially in drought-related biological processes. It also provides a direction for developing new varieties of poplar with drought resistance. Overall, our results provided significant information for further functional analysis of PtrHB genes in poplar and demonstrated that PtrHB3 is a dominant gene regulating tolerance to water stress treatment in poplar seedlings.

Case Report: Cutaneous Pleomorphic Lymphangiosarcoma in a Dog Exhibiting Features of Human Composite Hemangioendothelioma

Background: Angiosarcomas are a broad category of vascular origin neoplasms that are poorly characterized in veterinary species. Lymphangiosarcoma (LAS) is an uncommon type of angiosarcoma reported in humans and canines arising from lymphatic endothelium. LAS can be differentiated from other angiosarcomas in dogs based on expression of Prospero-related homeobox gene-1 (PROX-1) or lymphatic vessel endothelial receptor-1 (LYVE-1). Composite hemangioendothelioma (CHE) is a rare angiosarcoma subtype described in people and characterized by a variable biologic behavior and infrequent metastasis. This variant of angiosarcoma histologically combines features of retiform hemangioendothelioma and epithelioid hemangioendothelioma. Information regarding the cytologic and histopathologic appearance and clinical course of dogs with vascular tumors that exhibit features of CHE are unknown. Here, we report a case of pleomorphic LAS with features of CHE arising in a dog and treated with surgery and adjuvant chemotherapy.Case presentation: A 10-year-old intact male Labrador retriever presented with an approximately 6-cm-diameter cutaneous mass caudal to the left elbow that was progressively growing over 1.5 years. On physical examination, palpable extensions were identified coursing proximally over the triceps with concurrent loco-regional peripheral lymphadenopathy. Fine needle aspirates (FNA) and cytologic assessment of the cutaneous mass, left prescapular, and accessory axillary lymph nodes reported that this appeared to be a metastatic epithelial neoplasm, although a mixed carcinoma or collision tumor could not be excluded. An incisional biopsy of the mass was submitted for histopathology and was consistent with a well-differentiated angiosarcoma with features of CHE. The neoplasm expressed vimentin, CD31, von Willebrand factor (vWf), and PROX-1, supporting the diagnosis of LAS. Complete staging was performed, and no additional metastatic lesions were identified. Left forelimb amputation and lymph node removal were performed. Based on the diagnosis of metastatic LAS, doxorubicin chemotherapy was administered. 7 months post-amputation, the tumor recurred at the amputation site without evidence of metastatic disease.Conclusion: This report describes a malignant, locally aggressive lymphatic origin vascular tumor in a dog, with features consistent with descriptions of CHE in humans. Cytologic features in this case were discordant with its true mesenchymal etiology, obfuscating diagnosis. The morphologic features of the mesenchymal neoplastic population and immunohistochemistry (IHC) labeling ultimately supported a diagnosis of pleomorphic LAS with features of CHE.