Role of homeobox genes in cancer: immune system interactions, long non-coding RNAs, and tumor progression.

The intricate interplay between Homeobox genes, long non-coding RNAs (lncRNAs), and the development of malignancies represents a rapidly expanding area of research. Specific discernible lncRNAs have been discovered to adeptly regulate HOX gene expression in the context of cancer, providing fresh insights into the molecular mechanisms that govern cancer development and progression. An in-depth comprehension of these intricate associations may pave the way for innovative therapeutic strategies in cancer treatment. The HOX gene family is garnering increasing attention due to its involvement in immune system regulation, interaction with long non-coding RNAs, and tumor progression. Although initially recognized for its crucial role in embryonic development, this comprehensive exploration of the world of HOX genes contributes to our understanding of their diverse functions, potentially leading to immunology, developmental biology, and cancer research discoveries. Thus, the primary objective of this review is to delve into these aspects of HOX gene biology in greater detail, shedding light on their complex functions and potential therapeutic applications.

40†years of the homeobox: mechanisms of Hox spatial-temporal collinearity in vertebrates.

Animal body plans are established during embryonic development by the Hox genes. This patterning process relies on the differential expression of Hox genes along the head-to-tail axis. Hox spatial collinearity refers to the relationship between the organization of Hox genes in clusters and the differential Hox expression, whereby the relative order of the Hox genes within a cluster mirrors the spatial sequence of expression in the developing embryo. In vertebrates, the cluster organization is also associated with the timing of Hox activation, which harmonizes Hox expression with the progressive emergence of axial tissues. Thereby, in vertebrates, Hox temporal collinearity is intimately linked to Hox spatial collinearity. Understanding the mechanisms contributing to Hox temporal and spatial collinearity is thus key to the comprehension of vertebrate patterning. Here, we provide an overview of the main discoveries pertaining to the mechanisms of Hox spatial-temporal collinearity.

A taxon-restricted duplicate of Iroquois3 is required for patterning the spider waist.

The chelicerate body plan is distinguished from other arthropod groups by its division of segments into 2 tagmata: the anterior prosoma ("cephalothorax") and the posterior opisthosoma ("abdomen"). Little is understood about the genetic mechanisms that establish the prosomal-opisthosomal (PO) boundary. To discover these mechanisms, we created high-quality genomic resources for the large-bodied spider Aphonopelma hentzi. We sequenced specific territories along the antero-posterior axis of developing embryos and applied differential gene expression analyses to identify putative regulators of regional identity. After bioinformatic screening for candidate genes that were consistently highly expressed in only 1 tagma (either the prosoma or the opisthosoma), we validated the function of highly ranked candidates in the tractable spider model Parasteatoda tepidariorum. Here, we show that an arthropod homolog of the Iroquois complex of homeobox genes is required for proper formation of the boundary between arachnid tagmata. The function of this homolog had not been previously characterized, because it was lost in the common ancestor of Pancrustacea, precluding its investigation in well-studied insect model organisms. Knockdown of the spider copy of this gene, which we designate as waist-less, in P. tepidariorum resulted in embryos with defects in the PO boundary, incurring discontinuous spider germ bands. We show that waist-less is required for proper specification of the segments that span the prosoma-opisthosoma boundary, which in adult spiders corresponds to the narrowed pedicel. Our results demonstrate the requirement of an ancient, taxon-restricted paralog for the establishment of the tagmatic boundary that defines Chelicerata.

Effects of HOX family regulator-mediated modification patterns and immunity characteristics on tumor-associated cell type in endometrial cancer.

Endometrial cancer (UCEC) is one of three major malignant tumors in women. The HOX gene regulates tumor development. However, the potential roles of HOX in the expression mechanism of multiple cell types and in the development and progression of tumor microenvironment (TME) cell infiltration in UCEC remain unknown. In this study, we utilized both the The Cancer Genome Atlas (TCGA) database and International Cancer Genome Consortium (ICGC) database to analyze transcriptome data of 529 patients with UCEC based on 39 HOX genes, combing clinical information, we discovered HOX gene were a pivotal factor in the development and progression of UCEC and in the formation of TME diversity and complexity. Here, a new scoring system was developed to quantify individual HOX patterns in UCEC. Our study found that patients in the low HOX score group had abundant anti-tumor immune cell infiltration, good tumor differentiation, and better prognoses. In contrast, a high HOX score was associated with blockade of immune checkpoints, which enhances the response to immunotherapy. The Real-Time quantitative PCR (RT-qPCR) and Immunohistochemistry (IHC) exhibited a higher expression of the HOX gene in the tumor patients. We revealed that the significant upregulation of the HOX gene in the epithelial cells can activate signaling pathway associated with tumour invasion and metastasis through single-cell RNA sequencing (scRNA-seq), such as nucleotide metabolic proce and so on. Finally, a risk prognostic model established by the positive relationship between HOX scores and cancer-associated fibroblasts (CAFs) can predict the prognosis of individual patients by scRNA-seq and transcriptome data sets. In sum, HOX gene may serve as a potential biomarker for the diagnosis and prediction of UCEC and to develop more effective therapeutic strategies.

Irreducible Complexity of Hox Gene: Path to the Canonical Function of the Hox Cluster.

The evolution of major taxa is often associated with the emergence of new gene families. In all multicellular animals except sponges and comb jellies, the genomes contain Hox genes, which are crucial regulators of development. The canonical function of Hox genes involves colinear patterning of body parts in bilateral animals. This general function is implemented through complex, precisely coordinated mechanisms, not all of which are evolutionarily conserved and fully understood. We suggest that the emergence of this regulatory complexity was preceded by a stage of cooperation between more ancient morphogenetic programs or their individual elements. Footprints of these programs may be present in modern animals to execute non-canonical Hox functions. Non-canonical functions of Hox genes are involved in maintaining terminal nerve cell specificity, autophagy, oogenesis, pre-gastrulation embryogenesis, vertical signaling, and a number of general biological processes. These functions are realized by the basic properties of homeodomain protein and could have triggered the evolution of ParaHoxozoa and Nephrozoa subsequently. Some of these non-canonical Hox functions are discussed in our review.

Inhibition of ERN1 affects the expression of TGIF1 and other homeobox gene expressions in U87MG glioblastoma cells.

BACKGROUND: The ERN1 (endoplasmic reticulum to nucleus signaling 1) pathway plays an important role in the regulation of gene expression in glioblastoma, but molecular mechanism has not yet been fully elucidated. The aim of this study was to evaluate the relative relevance of ERN1 activity as a kinase in comparison to its endoribonuclease activity in the regulation of homeobox gene expression. METHODS: Two sublines of U87MG glioblastoma cells with different ways of ERN1 inhibition were used: dnERN1 (overexpressed transgene without protein kinase and endoribonuclease) and dnrERN1 (overexpressed transgene with mutation in endoribonuclease). ERN1 suppression was also done using siRNA for ERN1. Silencing of XBP1 mRNA by specific siRNA was used for suppression of ERN1 endoribonuclease function mediated by XBP1s. The expression levels of homeobox genes and microRNAs were evaluated by qPCR. RESULTS: The expression of TGIF1 and ZEB2 genes was downregulated in both types of glioblastoma cells with inhibition of ERN1 showing the ERN1 endoribonuclease-dependent mechanism of their regulation. However, the expression of PBX3 and PRPRX1 genes did not change significantly in dnrERN1 glioblastoma cells but was upregulated in dnERN1 cells indicating the dependence of these gene expressions on the ERN1 protein kinase. At the same time, the changes in PAX6 and PBXIP1 gene expressions introduced in glioblastoma cells by dnrERN1 and dnERN1 were different in direction and magnitude indicating the interaction of ERN1 protein kinase and endoribonuclease activities in regulation of these gene expressions. The impact of ERN1 and XBP1 silencing on the expression of studied homeobox genes is similar to that observed in dnERN1 and dnrERN1 glioblastoma cells, correspondingly. CONCLUSION: The expression of TGIF1 and other homeobox genes is dependent on the ern1 signaling pathways by diverse mechanisms because inhibition of ERN1 endoribonuclease and both ERN1 enzymatic activities had dissimilar impacts on the expression of most studied genes showing that ERN1 protein kinase plays an important role in controlling homeobox gene expression associated with glioblastoma cell invasion.

WUSCHEL-RELATED HOMEOBOX genes are crucial for normal vascular organization and wood formation in poplar.

Vascular cambium in tree species is a cylindrical domain of meristematic cells that are responsible for producing secondary xylem (also called wood) inward and secondary phloem outward. The poplar (Populus trichocarpa) WUSCHEL (WUS)-RELATED HOMEOBOX (WOX) family members, PtrWUSa and PtrWOX13b, were previously shown to be expressed in vascular cambium and differentiating xylem cells in poplar stems, but their functions remain unknown. Here, we investigated roles of PtrWUSa, PtrWOX13b and their close homologs in vascular organization and wood formation. Expression analysis showed that like PtrWUSa and PtrWOX13b, their close homologs, PtrWUSb, PtrWUS4a/b and PtrWOX13a/c, were also expressed in vascular cambium and differentiating xylem cells in poplar stems. PtrWUSa also exhibited a high level of expression in developing phloem fibers. Expression of PtrWUSa fused with the dominant EAR repression domain (PtrWUSa-DR) in transgenic poplar caused retarded growth of plants with twisted stems and curled leaves and a severe disruption of vascular organization. In PtrWUSa-DR stems, a drastic proliferation of cells occurred in the phloem region between vascular cambium and phloem fibers and they formed islands of ectopic vascular tissues or phloem fiber-like sclerenchyma cells. A similar proliferation of cells was also observed in PtrWUSa-DR leaf petioles and midveins. On the other hand, overexpression of PtrWOX4a-DR caused ectopic formation of vascular bundles in the cortical region, and overexpression of PtrWOX13a-DR and PtrWOX13b-DR led to a reduction in wood formation without affecting vascular organization in transgenic poplar plants. Together, these findings indicate crucial roles of PtrWUSa and PtrWOX13a/b in regulating vascular organization and wood formation, which furthers our understanding of the functions of WOX genes in regulating vascular cambium activity in tree species.

Teleost Hox code defines regional identities competent for the formation of dorsal and anal fins.

The dorsal and anal fins can vary widely in position and length along the anterior-posterior axis in teleost fishes. However, the molecular mechanisms underlying the diversification of these fins remain unknown. Here, we used genetic approaches in zebrafish and medaka, in which the relative positions of the dorsal and anal fins are opposite, to demonstrate the crucial role of hox genes in the patterning of the teleost posterior body, including the dorsal and anal fins. By the CRISPR-Cas9-induced frameshift mutations and positional cloning of spontaneous dorsalfinless medaka, we show that various hox mutants exhibit the absence of dorsal or anal fins, or a stepwise posterior extension of these fins, with vertebral abnormalities. Our results indicate that multiple hox genes, primarily from hoxc-related clusters, encompass the regions responsible for the dorsal and anal fin formation along the anterior-posterior axis. These results further suggest that shifts in the anterior boundaries of hox expression which vary among fish species, lead to diversification in the position and size of the dorsal and anal fins, similar to how modulations in Hox expression can alter the number of anatomically distinct vertebrae in tetrapods. Furthermore, we show that hox genes responsible for dorsal fin formation are different between zebrafish and medaka. Our results suggest that a novel mechanism has occurred during teleost evolution, in which the gene network responsible for fin formation might have switched to the regulation downstream of other hox genes, leading to the remarkable diversity in the dorsal fin position.

Homeobox and Polycomb target gene methylation in human solid tumors.

DNA methylation is an epigenetic mark that plays an important role in defining cancer phenotypes, with global hypomethylation and focal hypermethylation at CpG islands observed in tumors. These methylation marks can also be used to define tumor types and provide an avenue for biomarker identification. The homeobox gene class is one that has potential for this use, as well as other genes that are Polycomb Repressive Complex 2 targets. To begin to unravel this relationship, we performed a pan-cancer DNA methylation analysis using sixteen Illumina HM450k array datasets from TCGA, delving into cancer-specific qualities and commonalities between tumor types with a focus on homeobox genes. Our comparisons of tumor to normal samples suggest that homeobox genes commonly harbor significant hypermethylated differentially methylated regions. We identified two homeobox genes, HOXA3 and HOXD10, that are hypermethylated in all 16 cancer types. Furthermore, we identified several potential homeobox gene biomarkers from our analysis that are uniquely methylated in only one tumor type and that could be used as screening tools in the future. Overall, our study demonstrates unique patterns of DNA methylation in multiple tumor types and expands on the interplay between the homeobox gene class and oncogenesis.

The Hox code responsible for the patterning of the anterior vertebrae in zebrafish.

The vertebral column is a characteristic structure of vertebrates. Genetic studies in mice have shown that Hox-mediated patterning plays a key role in specifying discrete anatomical regions of the vertebral column. Expression pattern analyses in several vertebrate embryos have provided correlative evidence that the anterior boundaries of Hox expression coincide with distinct anatomical vertebrae. However, because functional analyses have been limited to mice, it remains unclear which Hox genes actually function in vertebral patterning in other vertebrates. In this study, various zebrafish Hox mutants were generated for loss-of-function phenotypic analysis to functionally decipher the Hox code responsible for the zebrafish anterior vertebrae between the occipital and thoracic vertebrae. We found that Hox genes in HoxB- and HoxC-related clusters participate in regulating the morphology of the zebrafish anterior vertebrae. In addition, medaka hoxc6a was found to be responsible for anterior vertebral identity, as in zebrafish. Based on phenotypic similarities with Hoxc6 knockout mice, our results suggest that the Hox patterning system, including at least Hoxc6, may have been functionally established in the vertebral patterning of the common ancestor of ray-finned and lobe-finned fishes.

Hox genes and patterning the vertebrate body.

The diversity of vertebrate body plans is dizzying, yet stunning for the many things they have in common. Vertebrates have inhabited virtually every part of the earth from its coldest to warmest climates. They locomote by swimming, flying, walking, slithering, or climbing, or combinations of these behaviors. And they exist in many different sizes, from the smallest of frogs, fish and lizards to giraffes, elephants, and blue whales. Despite these differences, vertebrates follow a remarkably similar blueprint for the establishment of their body plan. Within the relatively small amount of time required to complete gastrulation, the process through which the three germ layers, ectoderm, mesoderm, and endoderm are created, the embryo also generates its body axis and is simultaneously patterned. For the length of this axis, the genes that distinguish the neck from the rib cage or the trunk from the sacrum are the Hox genes. In vertebrates, there was evolutionary pressure to maintain this set of genes in the organism. Over the past decades, much has been learned regarding the regulatory mechanisms that ensure the appropriate expression of these genes along the main body axes. Genetic functions continue to be explored though much has been learned. Much less has been discerned on the identity of co-factors used by Hox proteins for the specificity of transcriptional regulation or what downstream targets and pathways are critical for patterning events, though there are notable exceptions. Current work in the field is demonstrating that Hox genes continue to function in many organs long after directing early patterning events. It is hopeful continued research will shed light on remaining questions regarding mechanisms used by this important and conserved set of transcriptional regulators.

The Evolution of the WUSCHEL-Related Homeobox Gene Family in Dendrobium Species and Its Role in Sex Organ Development in D. chrysotoxum.

The WUSCHEL-related homeobox (WOX) transcription factor plays a vital role in stem cell maintenance and organ morphogenesis, which are essential processes for plant growth and development. Dendrobium chrysotoxum, D. huoshanense, and D. nobile are valued for their ornamental and medicinal properties. However, the specific functions of the WOX gene family in Dendrobium species are not well understood. In our study, a total of 30 WOX genes were present in the genomes of the three Dendrobium species (nine DchWOXs, 11 DhuWOXs, and ten DnoWOXs). These 30 WOXs were clustered into ancient clades, intermediate clades, and WUS/modern clades. All 30 WOXs contained a conserved homeodomain, and the conserved motifs and gene structures were similar among WOXs belonging to the same branch. D. chrysotoxum and D. huoshanense had one pair of fragment duplication genes and one pair of tandem duplication genes, respectively; D. nobile had two pairs of fragment duplication genes. The cis-acting regulatory elements (CREs) in the WOX promoter region were mainly enriched in the light response, stress response, and plant growth and development regulation. The expression pattern and RT-qPCR analysis revealed that the WOXs were involved in regulating the floral organ development of D. chrysotoxum. Among them, the high expression of DchWOX3 suggests that it might be involved in controlling lip development, whereas DchWOX5 might be involved in controlling ovary development. In conclusion, this work lays the groundwork for an in-depth investigation into the functions of WOX genes and their regulatory role in Dendrobium species' floral organ development.

Hox gene-specific cellular targeting using split intein Trojan exons.

The Trojan exon method, which makes use of intronically inserted T2A-Gal4 cassettes, has been widely used in Drosophila to create thousands of gene-specific Gal4 driver lines. These dual-purpose lines provide genetic access to specific cell types based on their expression of a native gene while simultaneously mutating one allele of the gene to enable loss-of-function analysis in homozygous animals. While this dual use is often an advantage, the truncation mutations produced by Trojan exons are sometimes deleterious in heterozygotes, perhaps by creating translation products with dominant negative effects. Such mutagenic effects can cause developmental lethality as has been observed with genes encoding essential transcription factors. Given the importance of transcription factors in specifying cell type, alternative techniques for generating specific Gal4 lines that target them are required. Here, we introduce a modified Trojan exon method that retains the targeting fidelity and plug-and-play modularity of the original method but mitigates its mutagenic effects by exploiting the self-splicing capabilities of split inteins. "Split Intein Trojan exons" (siTrojans) ensure that the two truncation products generated from the interrupted allele of the native gene are trans-spliced to create a full-length native protein. We demonstrate the efficacy of siTrojans by generating a comprehensive toolkit of Gal4 and Split Gal4 lines for the segmentally expressed Hox transcription factors and illustrate their use in neural circuit mapping by targeting neurons according to their position along the anterior-posterior axis. Both the method and the Hox gene-specific toolkit introduced here should be broadly useful.

Role of HOX genes in cancer progression and their therapeutical aspects.

HOX genes constitute a family of evolutionarily conserved transcription factors that play pivotal roles in embryonic development, tissue patterning, and cell differentiation. These genes are essential for the precise spatial and temporal control of body axis formation in vertebrates. In addition to their developmental functions, HOX genes have garnered significant attention for their involvement in various diseases, including cancer. Deregulation of HOX gene expression has been observed in numerous malignancies, where they can influence tumorigenesis, progression, and therapeutic responses. This review provides an overview of the diverse roles of HOX genes in development, disease, and potential therapeutic targets, highlighting their significance in understanding biological processes and their potential clinical implications.

HOXB9 promotes laryngeal squamous cell carcinoma progression by upregulating MMP12.

Transcriptional factor HOXB9, a part of the HOX gene family, plays a crucial role in the development of diverse cancer types. This study aimed to elucidate the regulatory mechanism of HOXB9 on the proliferation and invasion of laryngeal squamous cell carcinoma (LSCC) cells to provide guidance for the development and prognosis of LSCC. The CRISPR/Cas9 method was employed in LSCC cell lines to knock out the HOXB9 gene and validate its effects on the proliferation, migration, invasion, and regulation of LSCC cells. CCK-8 and flow cytometry were used to detect cell viability and proliferation; Tunnel was used to detect cell apoptosis, and transwell was used to detect cell migration and invasion. The effect of HOXB9 on tumor growth was tested in nude mice. The downstream target genes regulated by HOXB9 were screened by microarray analysis and verified by Western blotting, immunohistochemistry, chromatin immunoprecipitation, and double-luciferase reporter assays. The current research investigated molecular pathways governed by HOXB9 in the development of LSCC. Additionally, both laboratory- and living-organism-based investigations revealed that disrupting the HOXB9 gene through the CRISPR/CAS9 mechanism restrained cellular growth, movement, and infiltration, while enhancing cellular apoptosis. Detailed analyses of LSCC cell strains and human LSCC samples revealed that HOXB9 promoted LSCC progression by directly elevating the transcriptional activity of MMP12. HOXB9 could influence changes in LSCC cell functions, and the mechanism of action might be exerted through its downstream target gene, MMP12.

Homeobox gene-encoded transcription factors in development and mature circadian function of the rodent pineal gland.

Homeobox genes encode transcription factors that are widely known to control developmental processes. This is also the case in the pineal gland, a neuroendocrine brain structure devoted to nighttime synthesis of the hormone melatonin. Thus, in accordance with high prenatal gene expression, knockout studies have identified a specific set of homeobox genes that are essential for development of the pineal gland. However, as a special feature of the pineal gland, homeobox gene expression persists into adulthood, and gene product abundance exhibits 24 h circadian rhythms. Recent lines of evidence show that some homeobox genes even control expression of enzymes catalyzing melatonin synthesis. We here review current knowledge of homeobox genes in the rodent pineal gland and suggest a model for dual functions of homeobox gene-encoded transcription factors in developmental and circadian mature neuroendocrine function.

HOXA9 gene inhibits proliferation and differentiation and promotes apoptosis of bovine preadipocytes.

BACKGROUND: Hox gene family is an important transcription factor that regulates cell process, and plays a role in the process of adipocytes differentiation and fat deposition. Previous transcriptome sequencing studies have indicated that the Homeobox A9 gene (HOXA9) is a candidate gene for regulating the process of bovine lipid metabolism, but the function and specific mechanism of action remain unclear. Therefore, this study aims to explore the role of HOXA9 in the proliferation, differentiation and apoptosis of bovine preadipocytes through gain-of-function and lose-of-function. RESULT: It found HOXA9 highly expressed in bovine adipose tissue, and its expression level changed significantly during adipocytes differentiation process. It gave a hint that HOXA9 may be involved in the process of bovine lipid metabolism. The results of HOXA9 gain-of-function experiments indicated that HOXA9 appeared to act as a negative regulator not only in the differentiation but also in the proliferation of bovine preadipocytes, which is mainly reflected that overexpression of HOXA9 down-regulate the mRNA and protein expression level of PPARγ, CEBPα and FABP4 (P < 0.05). The mRNA expression level of CDK1, CDK2, PCNA, CCNA2, CCNB1, CCND1 and CCNE2, as well as the protein expression of CDK2 also significantly decreased. The decrease of lipid droplets content was the main characteristic of the phenotype (P < 0.01), which further supported the evidence that HOXA9 was a negative regulator of preadipocytes differentiation. The decrease of cell proliferation rate and EdU positive rate, as well as the limitation of transition of preadipocytes from G0/G1 phase to S phase also provided evidence for the inhibition of proliferation. Apart from this above, we noted an interesting phenomenon that overexpression of HOXA9 showed in a significant upregulation of both mRNA and protein level of apoptosis markers, accompanied by a significant increase in cell apoptosis rate. These data led us not to refute the fact that HOXA9 played an active regulatory role in apoptosis. HOXA9 loss-of-function experiments, however, yielded the opposite results. Considering that HOXA9 acts as a transcription factor, we predicted its target genes. Dual luciferase reporter assay system indicated that overexpression of HOXA9 inhibits activity of PCNA promoter. CONCLUSION: Taken together, we demonstrated for the first time that HOXA9 played a role as a negative regulatory factor in the differentiation and proliferation of preadipocytes, but played a positive regulatory role in apoptosis, and it may play a regulatory role by targeting PCNA. This study provides basic data for further exploring the regulatory network of intramuscular fat deposition in bovine.

Homeobox B2 promotes malignant behavior and contributes to the radioresistance of nasopharyngeal carcinoma by regulating forkhead box protein O1.

Background: Nasopharyngeal carcinoma (NPC) is an epithelial tumor of the head and neck with heterogeneous racial and geographical distributions. Homeobox B2 (HOXB2) is a tumor promoter in many cancers. However, the biological role of HOXB2 in NPC has not been elucidated. Methods: Bioinformatics analysis was performed to identify the differentially expressed genes (DEGs) between samples of patients with radiosensitive and radioresistant NPC. qRT-PCR, western blotting and immunohistochemistry were used to detect the expression levels of the corresponding mRNA and proteins. Cell viability was detected by CCK-8 assay and colony-forming capability was evaluated using colony formation assays. Further, migration and invasion abilities were examined using wound-healing and transwell chamber assays, respectively. Cellular apoptosis after irradiation was assessed using flow cytometry and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining. Results: HOXB2 was identified as a potential regulator of radioresistance in NPC. Our in vitro results indicate that HOXB2 overexpression (HOXB2-OE) promoted malignant behaviors including invasion, migration, proliferation, and inhibited the irradiation-induced apoptosis of NPC cells. Consistent with these results, HOXB2 knockdown (HOXB2-sh) exhibited the opposite trends in these biological activities. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the DEGs were enriched in the FOXO signaling pathway. Mechanistically, western blotting showed that HOXB2-OE inhibited forkhead box protein O1 (FOXO1) expression in NPC cells. Thereafter, we transferred the FOXO1-OE plasmid to HOXB2-OE NPC cells and found that overexpression of FOXO1 reversed cell proliferation, migration, invasion, and radioresistance profiles promoted by HOXB2 overexpression. Conclusion: Our findings showed that HOXB2 acts as a tumor promoter in NPC, activating malignant behaviors and radioresistance of tumors via FOXO1 regulation. Moreover, the inactivation of HOXB2 or activation of FOXO1 are potential strategies to inhibit tumor progression and overcome radioresistance in NPC.

ERN1 knockdown modifies the hypoxic regulation of homeobox gene expression in U87MG glioblastoma cells.

OBJECTIVE.: Homeobox genes play an important role in health and disease including oncogenesis. The present investigation aimed to study ERN1-dependent hypoxic regulation of the expression of genes encoding homeobox proteins MEIS (zinc finger E-box binding homeobox 2) and LIM homeobox 1 family, SPAG4 (sperm associated antigen 4) and NKX3-1 (NK3 homeobox 1) in U87MG glioblastoma cells in response to inhibition of ERN1 (endoplasmic reticulum to nucleus signaling 1) for evaluation of their possible significance in the control of glioblastoma growth. METHODS.: The expression level of homeobox genes was studied in control (transfected by vector) and ERN1 knockdown U87MG glioblastoma cells under hypoxia induced by dimethyloxalylglycine (0.5 mM for 4 h) by quantitative polymerase chain reaction and normalized to ACTB. RESULTS.: It was found that hypoxia down-regulated the expression level of LHX2, LHX6, MEIS2, and NKX3-1 genes but up-regulated the expression level of MEIS1, LHX1, MEIS3, and SPAG4 genes in control glioblastoma cells. At the same time, ERN1 knockdown of glioblastoma cells significantly modified the sensitivity of all studied genes to a hypoxic condition. Thus, ERN1 knockdown of glioblastoma cells removed the effect of hypoxia on the expression of MEIS1 and LHX1 genes, but increased the sensitivity of MEIS2, LHX2, and LHX6 genes to hypoxia. However, the expression of MEIS3, NKX3-1, and SPAG4 genes had decreased sensitivity to hypoxia in ERN1 knockdown glioblastoma cells. Moreover, more pronounced changes under the conditions of ERN1 inhibition were detected for the pro-oncogenic gene SPAG4. CONCLUSION.: The results of the present study demonstrate that hypoxia affected the expression of homeobox genes MEIS1, MEIS2, MEIS3, LHX1, LHX2, LHX6, SPAG4, and NKX3-1 in U87MG glioblastoma cells in gene-specific manner and that the sensitivity of all studied genes to hypoxia condition is mediated by ERN1, the major pathway of the endoplasmic reticulum stress signaling, and possibly contributed to the control of glioblastoma growth. A fundamentally new results of this work is the establishment of the fact regarding the dependence of hypoxic regulation of SPAG4 gene expression on ER stress, in particular ERN1, which is associated with suppression of cell proliferation and tumor growth.

Generation of human excitatory forebrain neurons by cooperative binding of proneural NGN2 and homeobox factor EMX1.

Generation of defined neuronal subtypes from human pluripotent stem cells remains a challenge. The proneural factor NGN2 has been shown to overcome experimental variability observed by morphogen-guided differentiation and directly converts pluripotent stem cells into neurons, but their cellular heterogeneity has not been investigated yet. Here, we found that NGN2 reproducibly produces three different kinds of excitatory neurons characterized by partial coactivation of other neurotransmitter programs. We explored two principle approaches to achieve more precise specification: prepatterning the chromatin landscape that NGN2 is exposed to and combining NGN2 with region-specific transcription factors. Unexpectedly, the chromatin context of regionalized neural progenitors only mildly altered genomic NGN2 binding and its transcriptional response and did not affect neurotransmitter specification. In contrast, coexpression of region-specific homeobox factors such as EMX1 resulted in drastic redistribution of NGN2 including recruitment to homeobox targets and resulted in glutamatergic neurons with silenced nonglutamatergic programs. These results provide the molecular basis for a blueprint for improved strategies for generating a plethora of defined neuronal subpopulations from pluripotent stem cells for therapeutic or disease-modeling purposes.