HMG-box Domain Research Articles

Biallelic HMGXB4 loss-of-function variant causes intellectual disability, developmental delay, and dysmorphic features

BackgroundHMGXB4 (additionally known as HMG2L1) is a non-histone DNA-binding protein that contains a single HMG-box domain. HMGXB4 was originally described in Xenopus where it was seen to negatively regulate the Wnt/β-catenin signaling pathway. Materials and methodsIn this study, we conducted a genetic and clinical evaluation of a single family with three affected individuals suffering from intellectual disability (ID), global developmental delay (GDD) and dysmorphic facial features.Whole genome sequencing (WGS) and Sanger sequencing were performed on the affected individuals' DNA to identify genetic variations. Additionally, a reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to assess gene expression in both the affected and unaffected individuals in the family. ResultWGS identified a homozygous frameshift variant c.1193_1196del p. (Lys398Argfs × 25) in exon 5 of the HMGXB4 gene (OMIM 604702), which completely segregated the disease phenotype in the family. Furthermore, RT-qPCR revealed a substantial decrease in the HMGXB4 gene expression in the affected individuals as compared to the unaffected individuals of the family. ConclusionsThe current study is the first evidence linking a genetic variant in the HMGXB4 gene to ID, GDD, and dysmorphic facial features. Therefore, it is possible that HMGXB4 contributes significantly to developmental milestones and may be responsible for neurodevelopmental disorders in humans.

RNA Binding Properties of SOX Family Members.

SOX proteins are a family of transcription factors (TFs) that play critical functions in sex determination, neurogenesis, and chondrocyte differentiation, as well as cardiac, vascular, and lymphatic development. There are 20 SOX family members in humans, each sharing a 79-residue L-shaped high mobility group (HMG)-box domain that is responsible for DNA binding. SOX2 was recently shown to interact with long non-coding RNA and large-intergenic non-coding RNA to regulate embryonic stem cell and neuronal differentiation. The RNA binding region was shown to reside within the HMG-box domain; however, the structural details of this binding remain unclear. Here, we show that all SOX family members, except group H, interact with RNA. Our mutational experiments demonstrate that the disordered C-terminal region of the HMG-box domain plays an important role in RNA binding. Further, by determining a high-resolution structure of the HMG-box domain of the group H family member SOX30, we show that despite differences in RNA binding ability, SOX30 shares a very similar secondary structure with other SOX protein HMG-box domains. Together, our study provides insight into the interaction of SOX TFs with RNA.

Genome-Wide Identification and Expression Analysis of the High-Mobility Group B (HMGB) Gene Family in Plant Response to Abiotic Stress in Tomato.

High-mobility group B (HMGB) proteins are a class of non-histone proteins associated with eukaryotic chromatin and are known to regulate a variety of biological processes in plants. However, the functions of HMGB genes in tomato (Solanum lycopersicum) remain largely unexplored. Here, we identified 11 members of the HMGB family in tomato using BLAST. We employed genome-wide identification, gene structure analysis, domain conservation analysis, cis-acting element analysis, collinearity analysis, and qRT-PCR-based expression analysis to study these 11 genes. These genes were categorized into four groups based on their unique protein domain structures. Despite their structural diversity, all members contain the HMG-box domain, a characteristic feature of the HMG superfamily. Syntenic analysis suggested that tomato SlHMGBs have close evolutionary relationships with their homologs in other dicots. The promoter regions of SlHMGBs are enriched with numerous cis-elements related to plant growth and development, phytohormone responsiveness, and stress responsiveness. Furthermore, SlHMGB members exhibited distinct tissue-specific expression profiles, suggesting their potential roles in regulating various aspects of plant growth and development. Most SlHMGB genes respond to a variety of abiotic stresses, including salt, drought, heat, and cold. For instance, SlHMGB2 and SlHMGB4 showed positive responses to salt, drought, and cold stresses. SlHMGB1, SlHMGB3, and SlHMGB8 were involved in responses to two types of stress: SlHMGB1 responded to drought and heat, while SlHMGB3 and SlHMGB8 responded to salt and heat. SlHMGB6 and SlHMGB11 were solely regulated by drought and heat stress, respectively. Under various treatment conditions, the number of up-regulated genes significantly outnumbered the down-regulated genes, implying that the SlHMGB family may play a crucial role in mitigating abiotic stress in tomato. These findings lay a foundation for further dissecting the precise roles of SlHMGB genes.

PATH-02. NOVEL SOX10 INDEL MUTATIONS DRIVE SCHWANNOMAS THROUGH IMPAIRED TRANSACTIVATION OF MYELINATION GENE PROGRAMS

Abstract Schwannomas are common peripheral nerve sheath tumors that can cause severe morbidity due to their stereotypic intracranial and paraspinal locations. However, the molecular drivers responsible for a substantial subset remain unknown. Through genomic profiling of 96 schwannomas, we identified novel in-frame insertion/deletion mutations in the SOX10 gene in 29% of sporadic tumors which uniformly lacked alterations in known nerve sheath tumor genes including NF1, NF2, LZTR1, and SMARCB1. These indel mutations clustered at the C-terminal end of the HMG-box domain, the DNA binding motif of the SOX10 transcription factor critical for Schwann cell differentiation. Schwannomas arising from non-vestibular cranial nerves (e.g. facial, trigeminal, vagus) were highly enriched for SOX10 mutations, whereas the vast majority of vestibular schwannomas had NF2 inactivation. DNA methylation profiling revealed that SOX10 mutant schwannomas clustered together with schwannomas harboring NF2 mutation or SH3PXD2A-HTRA1 fusion, indicating that SOX10 indel mutations define a novel schwannoma molecular subtype and not a distinct nerve sheath tumor entity. To investigate the mechanism by which SOX10 indel mutations promote schwannoma development, we stably transduced human fetal glial cells with either wildtype SOX10 or two different tumor-derived insertion mutants. RNA sequencing revealed that both SOX10 indel mutants, in contrast to wildtype SOX10, failed to activate expression of myelination and glial differentiation gene programs, including PMP2 which encodes the major peripheral myelin protein in mature Schwann cells. Electrophoretic mobility shift assay revealed that the indel mutants retained their DNA binding capacity to the SOX10 binding site in the PMP2 promoter, while a luciferase reporter assay revealed transactivation at the PMP2 promoter by wildtype SOX10 but not by the mutants. In summary, we identified a new recurrent genetic event in 29% of sporadic schwannomas that we speculate drives schwannoma development through impaired transactivation of myelination gene expression programs causing blocked differentiation of immature Schwann cells.

Structural analysis of the Candida albicans mitochondrial DNA maintenance factor Gcf1p reveals a dynamic DNA-bridging mechanism.

The compaction of mitochondrial DNA (mtDNA) is regulated by architectural HMG-box proteins whose limited cross-species similarity suggests diverse underlying mechanisms. Viability of Candida albicans, a human antibiotic-resistant mucosal pathogen, is compromised by altering mtDNA regulators. Among them, there is the mtDNA maintenance factor Gcf1p, which differs in sequence and structure from its human and Saccharomyces cerevisiae counterparts, TFAM and Abf2p. Our crystallographic, biophysical, biochemical and computational analysis showed that Gcf1p forms dynamic protein/DNA multimers by a combined action of an N-terminal unstructured tail and a long helix. Furthermore, an HMG-box domain canonically binds the minor groove and dramatically bends the DNA while, unprecedentedly, a second HMG-box binds the major groove without imposing distortions. This architectural protein thus uses its multiple domains to bridge co-aligned DNA segments without altering the DNA topology, revealing a new mechanism of mtDNA condensation.

Abstract 3721: The C1 domain of capicua has functional importance in human CIC-DUX4

Abstract Genomic rearrangements between the HMG-box family transcriptional repressor capicua (CIC) and multiple partner genes result in fusion oncoproteins that activate transcription to drive sarcoma growth and progression. In the case of CIC-DUX4, the most common CIC-rearranged fusion oncoprotein, the fusion often retains over 90% of the wild type CIC protein structure including the HMG box and C-terminal C1 domain. Structure-function studies in Drosophila have suggested that the C1 domain of CIC, a non-HMG box DNA binding domain, is important for the transactivating capacity of a synthetic fly-human CIC-DUX4. Consistent with these findings, we performed a retrospective analysis of over 70 breakpoints described from CIC-DUX4 human derived tumors, which revealed that the C1 domain is structurally conserved within the context of the CIC-DUX4 fusion. To further understand the functional role of the C1 domain in human CIC-DUX4 we generated human CIC-DUX4 C1 domain and HMG-box deletion (alone or in combination) mutant constructs and performed a series of studies to test the functional impact on CIC-DUX4 protein levels, subcellular localization, and transcriptional activity. Through these analyses we have found that deletion of the C1 domain does not impact CIC-DUX4 oncoprotein expression or localization but does dramatically reduce target gene expression. In ongoing in vivo studies, we aim to evaluate the significance of the C1 domain in inducing CIC-DUX4 driven tumorigenesis. Collectively, our findings validate the functional importance of the C1 domain in human CIC-DUX4 and to our knowledge provide the first harmonized analysis of breakpoint locations in CIC-rearranged sarcoma. Citation Format: Cuyler Luck, Kyle A. Jacobs, Ross A. Okimoto. The C1 domain of capicua has functional importance in human CIC-DUX4. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3721.

The involvement of an HMG-box gene in germ cell genesis in Pyropia haitanensis

High mobility group (HMG)-box proteins play important roles in sex differentiation and determination of fungi, brown algae, and vertebrates. So far, the HMG-box genes from the red algal species have not been explored, and their function in this lineage is still unknown. In this study, based on homologous searching approach, solo member of HMG-box gene (PhHMG) was identified from the genomes of Pyropia haitanensis, the number of which was far less than that of brown algae, green algae, and higher plants. The PhHMG protein contains three HMG-box domains and is localized in the nucleus and cell membrane region according to subcellular localization analysis. Further gene expression analysis indicated that PhHMG may involve in the germ cells genesis. As no germ cells (carpogonia or spermatangium) appeared in 30-days aged blades of P. haitanensis, PhHMG was not expressed or slightly expressed. Accompanied by the gradually appeared germ cells, the expression level of PhHMG gradually increased, with no difference between female and male strains. In addition, the 40-days aged blades of WT-10 strain (♀) had more carpogonia at the apical parts, followed by the middle parts, and almost no carpogonia at the basal parts, and the expression of PhHMG showed a consistent trend of apical > middle > basal. This study has fundamental reference value in revealing the reproductive regulation mechanism and developing molecular assisted breeding of P. haitanensis.

HMGN2 represses gene transcription via interaction with transcription factors Lef-1 and Pitx2 during amelogenesis

The chromatin-associated high mobility group protein N2 (HMGN2) cofactor regulates transcription factor activity through both chromatin and protein interactions. Hmgn2 expression is known to be developmentally regulated, but the post-transcriptional mechanisms that regulate Hmgn2 expression and its precise roles in tooth development remain unclear. Here, we demonstrate that HMGN2 inhibits the activity of multiple transcription factors as a general mechanism to regulate early development. Bimolecular fluorescence complementation, pull-down, and coimmunoprecipitation assays show that HMGN2 interacts with the transcription factor Lef-1 through its HMG-box domain as well as with other early development transcription factors, Dlx2, FoxJ1, and Pitx2. Furthermore, EMSAs demonstrate that HMGN2 binding to Lef-1 inhibits its DNA-binding activity. We found that Pitx2 and Hmgn2 associate with H4K5ac and H3K4me2 chromatin marks in the proximal Dlx2 promoter, demonstrating Hmgn2 association with open chromatin. In addition, we demonstrate that microRNAs (miRs) mir-23a and miR-23b directly target Hmgn2, promoting transcriptional activation at several gene promoters, including the amelogenin promoter. In vivo, we found that decreased Hmgn2 expression correlates with increased miR-23 expression in craniofacial tissues as the murine embryo develops. Finally, we show that ablation of Hmgn2 in mice results in increased amelogenin expression because of increased Pitx2, Dlx2, Lef-1, and FoxJ1 transcriptional activity. Taken together, our results demonstrate both post-transcriptional regulation of Hmgn2 by miR-23a/b and post-translational regulation of gene expression by Hmgn2–transcription factor interactions. We conclude that HMGN2 regulates tooth development through its interaction with multiple transcription factors.

The C-Terminal Acidic Tail Modulates the Anticancer Properties of HMGB1.

Energy metabolism reprogramming was recently listed as a hallmark of cancer. In this process, the switch from pyruvate kinase isoenzyme type M1 to pyruvate kinase isoenzyme type M2 (PKM2) is believed to play a crucial role. Interestingly, the activity of the active form of PKM2 can efficiently be inhibited by the high-mobility group box 1 (HMGB1) protein, leading to a rapid blockage of glucose-dependent aerobic respiration and cancer cell death. HMGB1 is a member of the HMG protein family. It contains two DNA-binding HMG-box domains and an acidic C-terminal tail capable of positively or negatively modulating its biological properties. In this work, we report that the deletion of the C-terminal tail of HMGB1 increases its activity towards a large panel of cancer cells without affecting the viability of normal immortalized fibroblasts. Moreover, in silico analysis suggests that the truncated form of HMGB1 retains the capacity of the full-length protein to interact with PKM2. However, based on the capacity of the cells to circumvent oxidative phosphorylation inhibition, we were able to identify either a cytotoxic or cytostatic effect of the proteins. Together, our study provides new insights in the characterization of the anticancer activity of HMGB1.

Evolution, characterization and expression analysis of Sox gene family in rainbow trout (Oncorhynchus mykiss)

The Sox transcription factor family plays an important role in various biological processes such as animal sex determination and multiple organ development. We used online databases to analyze the gene structure, chemical characteristics, and evolutionary relationship of Sox family genes through bioinformatics, and we studied the expression profiles and regulatory mechanisms of Sox family genes. A total of 29 rainbow trout Sox genes were identified. The phylogenetic analysis found that Sox genes of rainbow trout were clustered in seven subfamilies (B1, B2, C, D, E, F and H), and the gene structure of each subfamily was relatively conserved. Furthermore, Sox1, Sox4, Sox6, Sox8, Sox9, Sox11, Sox17, Sox18, and Sox19 developed into two copies, which might be the result of teleost fish-specific genome replication. Multiple HMG box domain alignments indicated that the motifs for all Sox sequences are conserved. Gene expression studies reveal that Sox expression is tissue-specific and that multiple Sox genes are involved in rainbow trout gonad and central nervous system development. Our study provides valuable information on the evolution of teleosts, and will also help to further research the functional characteristics of Sox genes.

Exploration of naphthoquinone analogs in targeting the TCF-DNA interaction to inhibit the Wnt/β-catenin signaling pathway

The Wnt/β-catenin signaling pathway plays extensive roles in cancer initiation, proliferation, and development, and has been implicated in the regulation of stem cells in the intestinal crypt, widely accepted as responsible for colorectal cancer (CRC) origination. This pathway has been a target of interest for many years for chemotherapeutic development of CRC due to its implication in most cases. Previously, a series of naphthoquinone analogs have been identified to inhibit the Wnt/β-catenin. It was postulated that these compounds exhibit their inhibitory activity via binding to β-catenin at the β-catenin/TCF4 interaction interface. In this study, we aimed to further define the critical pharmacophore for these compounds and verify their mechanisms of action for their abilities to inhibit the Wnt/β-catenin signaling pathway. Interestingly, our data suggested two of the compounds, compounds 3 and 6, may potently inhibit the Wnt/β-catenin signaling pathway via inhibition of the TCF4/DNA interaction, a novel finding compared to previous studies on these compounds. Our computational studies suggested that the compounds bound within the DNA binding HMG-box domain of TCF4 to elicit their inhibitory action. These compounds inhibited Wnt signaling in a dose dependent manner, suppressed Wnt direct target genes and demonstrated unforeseen degradation of the TCF4 protein. Thus, this study revealed a potentially novel mechanism of action of the chloro-naphthoquinone as possibly a multi-targeting scaffold, which warrants further investigation in future drug discovery on the ‘undruggable” TCF proteins and an aberrantly activated Wnt/β-catenin signaling pathway.

Circular RNA EGLN3 silencing represses renal cell carcinoma progression through the miR-1224-3p/HMGXB3 axis

BackgroundRenal cell carcinoma (RCC) is a common tumor of the urinary system, and its global incidence is increasing annually. Circular RNAs (circRNAs) are involved in RCC tumorigenesis; however, the role of circ-EGLN3 (hsa_circ_0031594) derived from the Egl nine homolog 3 (EGLN3) gene in RCC remains undetermined. MethodsCirc-EGNL3 expression was examined before and after RNase R and actinomycin treatments in RCC cells and tissues. Cell proliferation, migration, and invasion were assessed using the CCK-8 assay, EdU staining, and wound-healing and Transwell assays. The interactions between microRNA (miR)-1224-3p and circ-EGLN3, and between miR-1224-3p and HMG box domain containing 3 (HMGXB3) were predicted by bioinformatics analysis and validated by dual-luciferase reporter assay. ResultsCirc-EGLN3 was identified using RNase R and actinomycin treatments. Circ-EGLN3 was upregulated in RCC cells and tissues and correlated with poor overall survival. Silencing of circ-EGNL3 decreased RCC cell proliferation, migration, and invasion. Mechanistic studies indicated that circ-EGNL3 acts as a sponge for miR-1224-3p, which targeted HMGXB3. Circ-EGNL3 indirectly upregulated HMGXB3 by targeting miR-1224-3p, and overexpression of circ-EGLN3 reversed the repressive effects of miR-1224-3p on RCC. ConclusionCirc-EGLN3 regulated RCC progression through the miR-1224-3p/HMGXB3 axis, suggesting its potential as a therapeutic target.

HBD1 protein with a tandem repeat of two HMG-box domains is a DNA clip to organize chloroplast nucleoids in Chlamydomonas reinhardtii

Compaction of bulky DNA is a universal issue for all DNA-based life forms. Chloroplast nucleoids (chloroplast DNA-protein complexes) are critical for chloroplast DNA maintenance and transcription, thereby supporting photosynthesis, but their detailed structure remains enigmatic. Our proteomic analysis of chloroplast nucleoids of the green alga Chlamydomonas reinhardtii identified a protein (HBD1) with a tandem repeat of two DNA-binding high mobility group box (HMG-box) domains, which is structurally similar to major mitochondrial nucleoid proteins transcription factor A, mitochondrial (TFAM), and ARS binding factor 2 protein (Abf2p). Disruption of the HBD1 gene by CRISPR-Cas9-mediated genome editing resulted in the scattering of chloroplast nucleoids. This phenotype was complemented when intact HBD1 was reintroduced, whereas a truncated HBD1 with a single HMG-box domain failed to complement the phenotype. Furthermore, ectopic expression of HBD1 in the mitochondria of yeast Δabf2 mutant successfully complemented the defects, suggesting functional similarity between HBD1 and Abf2p. Furthermore, in vitro assays of HBD1, including the electrophoretic mobility shift assay and DNA origami/atomic force microscopy, showed that HBD1 is capable of introducing U-turns and cross-strand bridges, indicating that proteins with two HMG-box domains would function as DNA clips to compact DNA in both chloroplast and mitochondrial nucleoids.

HMGB1-like dorsal switch protein 1 of the mealworm, Tenebrio molitor, acts as a damage-associated molecular pattern.

High-mobility group box 1 (HMGB1) is a nuclear protein highly conserved in eukaryotes and ubiquitously expressed to regulate transcription and chromatin remodeling. Dorsal switch protein 1 (DSP1) is its insect homolog. A lepidopteran DSP1 acts as a damage-associated molecular pattern (DAMP) in response to immune challenge. The objective of this study was to determine the role of DAMP in the mealworm beetle, Tenebrio molitor, a coleopteran insect. DSP1 of T. molitor (Tm-DSP1) encodes 536 amino acids and shares sequence similarities with Homo sapiens HMGB1 (56.3%) and Spodoptera exigua DSP1 (59.2%). An antisera raised against S. exigua DSP1 was cross-reactive to Tm-DSP1. Like other insect DSPs, Tm-DSP1 has a relatively long N-terminal extension in addition to two conserved HMG box domains. It was expressed in all developmental stages of T. molitor and different larval tissues. Upon immune challenge, its expression level was upregulated. Its RNA interference (RNAi) treatment resulted in a significant reduction in immune responses measured by hemocyte nodule formation against bacterial infection. In addition, the induction of some antimicrobial peptide genes to the immune challenge was suppressed by its RNAi treatment. Interestingly, phospholipase A2 associated with eicosanoid biosynthesis was significantly suppressed in its catalytic activity by the RNAi treatment specific to Tm-DSP1 expression. Without any pathogen infection, injection of a lepidopteran DSP1 induced both cellular and humoral immune responses. These results suggest that Tm-DSP1 in T. molitor can act as a DAMP molecule and mediate immune responses upon immune challenge.

Genome wide identification, phylogeny, and synteny analysis of sox gene family in common carp (Cyprinus carpio)

Common carp (Cyprinus carpio) is a commercial fish species valuable for nutritious components and plays a vital role in human healthy nutrition. The SOX (SRY-related genes systematically characterized by a high-mobility group HMG-box) encoded important gene regulatory proteins, a family of transcription factors found in a broad range of animal taxa and extensively known for its contribution in multiple developmental processes including contribution in sex determination across phyla. In our current study, we initially accomplished a genome-wide analysis to report the SOX gene family in common carp fish based on available genomic sequences of zebrafish retrieved from gene repository databases, we focused on the global identification of the Sox gene family in Common carp among wide range of vertebrates and teleosts based on bioinformatics tools and techniques and explore the evolutionary relationships. In our results, a total of 27 SOX (high-mobility group HMG-box) domain genes were identified in the C. carp genome. The full length sequences of SOX genes ranging from 3496 (SOX6) to 924bp (SOX17b) which coded with putative proteins series from 307 to 509 amino acids and all gene having exon number expect SOX9 and SOX13. All the SOX proteins contained at least one conserved DNA-binding HMG-box domain and two (SOX7 and SOX18) were found C terminal. The Gene ontology revealed SOX proteins maximum involvement is in metabolic process 49.796 %, average in biological regulation 45.188 %, biosynthetic process (19.992 %), regulation of cellular process 39.68, 45.508 % organic substance metabolic process, multicellular organismal process 23.23 %,developmental process 21.74 %, system development 16.59 %, gene expression 16.05 % and 14.337 % of RNA metabolic process. Chromosomal location and syntanic analysis show all SOX gene are located on different chromosomes and apparently does not fallow the unique pattern. The maximum linkage of chromosome is (2) on Unplaced Scaffold region. Finally, our results provide important genomic suggestion for upcoming studies of biochemical, physiological, and phylogenetic understanding on SOX genes among teleost.

Structural basis for nuclear import selectivity of pioneer transcription factor SOX2

SOX (SRY-related HMG-box) transcription factors perform critical functions in development and cell differentiation. These roles depend on precise nuclear trafficking, with mutations in the nuclear targeting regions causing developmental diseases and a range of cancers. SOX protein nuclear localization is proposed to be mediated by two nuclear localization signals (NLSs) positioned within the extremities of the DNA-binding HMG-box domain and, although mutations within either cause disease, the mechanistic basis has remained unclear. Unexpectedly, we find here that these two distantly positioned NLSs of SOX2 contribute to a contiguous interface spanning 9 of the 10 ARM domains on the nuclear import adapter IMPα3. We identify key binding determinants and show this interface is critical for neural stem cell maintenance and for Drosophila development. Moreover, we identify a structural basis for the preference of SOX2 binding to IMPα3. In addition to defining the structural basis for SOX protein localization, these results provide a platform for understanding how mutations and post-translational modifications within these regions may modulate nuclear localization and result in clinical disease, and also how other proteins containing multiple NLSs may bind IMPα through an extended recognition interface.

Genome-wide identification and expression profile of the sox gene family in different tissues and during embryogenesis in the Pacific white shrimp (Litopenaeus vannamei).

Sox transcription factors play essential roles in a variety of critical physiological processes. Still, members of the sox gene family have not yet been genome-wide identified in shrimps. In this study, a total of five members of the sox gene family were identified from the genome of Pacific white shrimp Litopenaeus vannamei and classified into three subgroups based on the conserved HMG-box domain. Among them, three belong to the SoxB subgroup (one in B1 and two in B2), one in the SoxC subgroup, and one in the SoxE subgroup. The five sox genes had different sex-biased expression in some tissues. Sox21, soxB1, and sox14 had a higher expression in ovary than in testis. In comparison, sox4 had a male-biased specific expression in the gonad, hepatopancreas, gill, and eyestalk. There was no difference in soxE gene expression between testis and ovary. During embryonic development, the expression level of three sox genes (soxB1, sox21, and soxE) was higher in gastrulation stage compared to previous stages, declined in limb bud stage and then increased in intramembrane nauplius stage; the expression of sox4 was detected in blastula stage and continued to increase in the following two stages and then surged in intramembrane nauplius stage; the highest expression of sox14 was in the fertilized egg stage, and the expression level decreased with the development of the embryo. These results suggest that the shrimp sox gene family may be involved in gametogenesis, tridermogenesis, and neurogenesis.

Molecular characterization and expression profiles of transcription factor Sox gene family in Culter alburnus

Sox protein family is characterized by the presence of the conserved high-mobility group (HMG) box. Sox transcription factors are involved in diverse developmental process in animals, including sex-determination, organogenesis, embryogenesis, neurogenesis, and cell fate decision. In this study, 23 Sox genes were identified based on the Culter alburnus whole-genome sequence and categorized into six subfamilies according to the conserved HMG-box domain. The duplicates of four members revealed that Sox genes in the teleost fishes underwent significant expansion. Moreover, their expression pattern in gonad tissues were analyzed by RNA-seq and qRT-PCR, and Sox9b was determined as a key gene that was essential for testis development. This current study will provide new insight into the role of Sox gene family in fish sex determination and differentiation.

The novel Huntiella omanensis mating gene, MAT1-2-7, is essential for ascomatal maturation

Sexual reproduction is a highly conserved feature of the eukaryotes, yet sexual compatibility is determined by a wide variety of mechanisms. In ascomycete fungi, sexual development is controlled by genes at the mating type (MAT) locus that confer either MAT1-1 or MAT1-2 mating identity. Although the locus harbours, at minimum, a single gene, the individual MAT loci of certain species, including Huntiella omanensis, encode for two or more genes. The MAT1-2 idiomorph of H. omanensis is made up of MAT1-2-1, a primary MAT gene that is highly conserved in the Pezizomycotina and possesses a well-characterized DNA binding motif, the HMG-box domain. The idiomorph also harbours a novel secondary MAT gene, named MAT1-2-7, with no recognizable functional domains. In this study, we developed a transformation and CRISPR-Cas9-based genome editing protocol to characterize the MAT1-2-7 gene with respect to its function in mating. We have shown that MAT1-2-7 is essential for sexual reproduction and that isolates carrying the truncated MAT1-2-7 gene are incapable of ascomatal maturation and further sexual development. MAT1-2-7 was also shown to influence the vegetative radial growth rate of H. omanensis, illustrating the pleiotropic effects often associated with MAT genes.

Computer-Aided Discovery of Small Molecule Inhibitors of Thymocyte Selection-Associated High Mobility Group Box Protein (TOX) as Potential Therapeutics for Cutaneous T-Cell Lymphomas.

Cutaneous T-cell lymphomas (CTCL) are the most common primary lymphomas of the skin. We have previously identified thymocyte selection-associated high mobility group (HMG) box protein (TOX) as a promising drug target in CTCL; however, there are currently no small molecules able to directly inhibit TOX. We aimed to address this unmet opportunity by developing anti-TOX therapeutics with the use of computer-aided drug discovery methods. The available NMR-resolved structure of the TOX protein was used to model its DNA-binding HMG-box domain. To investigate the druggability of the corresponding protein–DNA interface on TOX, we performed a pilot virtual screening of 200,000 small molecules using in silico docking and identified ‘hot spots’ for drug-binding on the HMG-box domain. We then performed a large-scale virtual screening of 7.6 million drug-like compounds that were available from the ZINC15 database. As a result, a total of 140 top candidate compounds were selected for subsequent in vitro validation. Of those, 18 small molecules have been characterized as selective TOX inhibitors.