Box Domain Research Articles

The Molecular and Function Characterization of Porcine MID2.

Midline2 (MID2/TRIM1) is a member of the tripartite motif-containing (TRIM) family, which is involved in a wide range of cellular processes. However, fundamental studies on porcine MID2 (pMID2) are still lacking. In this study, we identified and characterized the full length MID2 gene of pig (Sus scrofa). The sequence alignment analysis results showed that pMID2 had an N-terminal RING zinc-finger domain, BBC domain, and C-terminal COS box, FN3 motif, and PRY-SPRY domain that were conserved and similar to those of other vertebrates. Furthermore, pMID2 had the highest expression levels in porcine lung and spleen. Serial deletion and site-directed mutagenesis showed that the putative nuclear factor-κB (NF-κB) binding site may be an essential transcription factor for regulating the transcription expression of pMID2. Furthermore, the immunofluorescence assay indicated that pMID2 presented in the cell membrane and cytoplasm. To further study the functions of pMID2, we identified and determined its potential ability to perceive poly (I:C) and IFN-α stimulation. Stimulation experiments showed pMID2 enhanced poly (I:C)-/IFN-α-induced JAK-STAT signaling pathway, indicating that pMID2 might participate in the immune responses. In conclusion, we systematically and comprehensively analyzed the characterizations and functions of pMID2, which provide valuable information to explore the pMID2 functions in innate immunity. Our findings not only enrich the current knowledge of MID2 in IFN signaling regulation but also offer the basis for future research of pig MID2 gene.

A Rab-bit hole: Rab40 GTPases as new regulators of the actin cytoskeleton and cell migration.

The regulation of machinery involved in cell migration is vital to the maintenance of proper organism function. When migration is dysregulated, a variety of phenotypes ranging from developmental disorders to cancer metastasis can occur. One of the primary structures involved in cell migration is the actin cytoskeleton. Actin assembly and disassembly form a variety of dynamic structures which provide the pushing and contractile forces necessary for cells to properly migrate. As such, actin dynamics are tightly regulated. Classically, the Rho family of GTPases are considered the major regulators of the actin cytoskeleton during cell migration. Together, this family establishes polarity in the migrating cell by stimulating the formation of various actin structures in specific cellular locations. However, while the Rho GTPases are acknowledged as the core machinery regulating actin dynamics and cell migration, a variety of other proteins have become established as modulators of actin structures and cell migration. One such group of proteins is the Rab40 family of GTPases, an evolutionarily and functionally unique family of Rabs. Rab40 originated as a single protein in the bilaterians and, through multiple duplication events, expanded to a four-protein family in higher primates. Furthermore, unlike other members of the Rab family, Rab40 proteins contain a C-terminally located suppressor of cytokine signaling (SOCS) box domain. Through the SOCS box, Rab40 proteins interact with Cullin5 to form an E3 ubiquitin ligase complex. As a member of this complex, Rab40 ubiquitinates its effectors, controlling their degradation, localization, and activation. Because substrates of the Rab40/Cullin5 complex can play a role in regulating actin structures and cell migration, the Rab40 family of proteins has recently emerged as unique modulators of cell migration machinery.

Karhunen-Loève expansion based on an analytical solution over a bounding box domain

This paper explores the accuracy and the efficiency of analytical solution of Fredholm integral equation to represent a random field on complex geometry. Because no analytical solution is available for arbitrary domains, it is proposed to use the analytical solution on simple bounding domains that enclose complex two- or three-dimensional geometries. It is a simple, accurate and robust approach for discretising a random field. The effect of the size of the bounding box on the resulting random field variance is investigated carefully and compared with the numerical solution given by the finite element method. The error variance is particularly localised near to the support domain boundary. Therefore, it is suggested to expand the bounding domain. This paper proposes a calibration of the correlation length to the ratio of the domain to maintain the convergence rate and the variance accuracy of the KLE without enlarging the stochastic dimension.

Seizure enhances SUMOylation and zinc-finger transcriptional repression in neuronal nuclei

A single episode of pilocarpine-induced status epilepticus can trigger the development of spontaneous recurrent seizures in a rodent model for epilepsy. The initial seizure-induced events in neuronal nuclei that lead to long-term changes in gene expression and cellular responses likely contribute toward epileptogenesis. Using a transgenic mouse model to specifically isolate excitatory neuronal nuclei, we profiled the seizure-induced nuclear proteome via tandem mass tag mass spectrometry and observed robust enrichment of nuclear proteins associated with the SUMOylation pathway. In parallel with nuclear proteome, we characterized nuclear gene expression by RNA sequencing which provided insights into seizure-driven transcriptional regulation and dynamics. Strikingly, we saw widespread downregulation of zinc-finger transcription factors, specifically proteins that harbor Krüppel-associated box (KRAB) domains. Our results provide a detailed snapshot of nuclear events induced by seizure activity and demonstrate a robust method for cell-type-specific nuclear profiling that can be applied to other cell types and models.

Leverage of nuclease-deficient CasX for preventing pathological angiogenesis

Gene editing with a CRISPR/Cas system is a novel potential strategy for treating human diseases. Pharmacological inhibition of phosphoinositide 3-kinase (PI3K) δ suppresses retinal angiogenesis in a mouse model of oxygen-induced retinopathy. Here we show that an innovative system of adeno-associated virus (AAV)-mediated CRISPR/nuclease-deficient (d)CasX fused with the Krueppel-associated box (KRAB) domain is leveraged to block (81.2%± 6.5%) invitro expression of p110δ, the catalytic subunit of PI3Kδ, encoded by Pik3cd. This CRISPR/dCasX-KRAB (4, 269bp) system is small enough to be fit into a single AAV vector. We then document that recombinant AAV serotype (rAAV)1 efficiently transduces vascular endothelial cells from pathologic retinal vessels, which show high expression of p110δ; furthermore, we demonstrate that blockade of retinal p110δ expression by intravitreally injected rAAV1-CRISPR/dCasX-KRAB targeting the Pik3cd promoter prevents (32.1%± 5.3%) retinal p110δ expression as well as pathological retinal angiogenesis in a mouse model of oxygen-induced retinopathy. These data establish a strong foundation for treating pathological angiogenesis by AAV-mediated CRISPR interference with p110δ expression.

<italic>Geminocystis urbisnovae</italic> sp. nov. (Chroococcales, Cyanobacteria): polyphasic description complemented with a survey of the family <italic>Geminocystaceae</italic>

Progress in phylogenomic analysis has led to a considerable re-evaluation of former cyanobacterial system, with many new taxa being established at different nomenclatural levels. The family <i>Geminocystaceae</i> is among cyanobacterial taxa recently described on the basis of polyphasic approach. Within this family, there are six genera: <i>Geminocystis</i>, <i>Cyanobacterium</i>, <i>Geminobacterium</i>, <i>Annamia</i>, <i>Picocyanobacterium</i>, and <i>Microcrocis</i>. The genus <i>Geminocystis</i> previously encompassed two species: <i>G. herdmanii </i>and <i>G. papuanica</i>. Herein, a new species <i>G. urbisnovae</i> was proposed under the provision of the International Code of Nomenclature for algae, fungi, and plants (ICN). Polyphasic analysis was performed for five strains from the CALU culture collection (St. Petersburg State University, Russian Federation), and they were assigned to the genus Geminocystis in accordance with high 16S rRNA gene similarity to existing species, as well as because of proximity to these species on the phylogenetic trees reconstructed with RaxML and Bayes methods. Plausibility of their assignment to a separate species of the genus <i>Geminocystis</i> was substantiated with smaller cell size; stenohaline freshwater ecotype; capability to complementary chromatic adaptation of second type (CA2); distinct 16S rRNA gene clustering; sequences and folding of D1−D1′ and B box domains of the 16S−23S internal transcribed spacer region. The second objective pursued by this communication was to provide a survey of the family <i>Geminocystaceae</i>. The overall assessment was that, despite attention of many researchers, this cyanobacterial family has been understudied and, especially in the case of the crucially important genus Cyanobacterium, taxonomically problematic.

A radiation box domain truncation scheme for the wave equation

Abstract We consider the wave equation in an unbounded domain and are interested in domain truncation methods. Our aim is to develop a numerical scheme that allows calculations for truncated waveguide geometries with periodic coefficient functions. The scheme is constructed with radiation boxes that are attached to the artificially introduced boundaries. A Dirichlet-to-Neumann operator $N$ is calculated in these radiation boxes. Efficiency of the scheme is obtained by calculating $N$ not with an iteration, but with a single run through the time interval. We observe speed-up factors of up to $20$ in comparison to calculations without domain truncation.

Isolation of FLOWERING LOCUS C and Preliminary Characterization in the Floral Transition of Xinjiang Precocious Walnut

Walnut (Juglans regia L.) plants typically flower after eight to ten years of juvenile growth. Precocious germplasm, also known as early-flowering or early-mature genotypes, have shortened juvenile phases of one to two years and are therefore crucial for enhancing breeding efficiency. However, such precocious germplasms are very limited. Here, we isolated and characterized the key flowering-time gene FLOWERING LOCUS C (FLC) in the precocious walnuts of the Xinjiang Uygur Autonomous Region. Sequence alignment showed that Juglans regia FLC (JrFLC)contained a conserved MINICHROMOSOME MAINTENANCE 1 (MCM1), AGAMOUS (AG), DEFICIENS (DEF), and SERUM RESPONSE FACTOR (SRF) (MADS)-box domain. Analysis of an FLC–green fluorescent protein (GFP) fusion protein revealed that JrFLC was localized to the nucleus. Gene expression analysis showed that JrFLC was specifically expressed during the bud dormancy stage of precocious walnut, and that expression levels gradually decreased as the ambient temperature warmed. Exogenous JrFLC overexpression in Arabidopsis thaliana delayed flowering and increased the total leaf number, suggesting a similar function of JrFLC as a floral repressor in walnut and in other plants. Together, these results showed that JrFLC played an important role in regulating the floral transition of Xinjiang precocious walnut. Further studies, including a detailed characterization of JrFLC, are expected to validate JrFLC as a strong target for genetic improvement in flowering time in walnut.

Pax4 in Health and Diabetes

Paired box 4 (Pax4) is a key transcription factor involved in the embryonic development of the pancreatic islets of Langerhans. Consisting of a conserved paired box domain and a homeodomain, this transcription factor plays an essential role in early endocrine progenitor cells, where it is necessary for cell-fate commitment towards the insulin-secreting β cell lineage. Knockout of Pax4 in animal models leads to the absence of β cells, which is accompanied by a significant increase in glucagon-producing α cells, and typically results in lethality within days after birth. Mutations in Pax4 that cause an impaired Pax4 function are associated with diabetes pathogenesis in humans. In adulthood, Pax4 expression is limited to a distinct subset of β cells that possess the ability to proliferate in response to heightened metabolic needs. Upregulation of Pax4 expression is known to promote β cell survival and proliferation. Additionally, ectopic expression of Pax4 in pancreatic islet α cells or δ cells has been found to generate functional β-like cells that can improve blood glucose regulation in experimental diabetes models. Therefore, Pax4 represents a promising therapeutic target for the protection and regeneration of β cells in the treatment of diabetes. The purpose of this review is to provide a thorough and up-to-date overview of the role of Pax4 in pancreatic β cells and its potential as a therapeutic target for diabetes.

A novel role of TRIM28 B box domain in L1 retrotransposition and ORF2p-mediated cDNA synthesis.

The long interspersed element 1 (LINE-1 or L1) integration is affected by many cellular factors through various mechanisms. Some of these factors are required for L1 amplification, while others either suppress or enhance specific steps during L1 propagation. Previously, TRIM28 has been identified to suppress transposable elements, including L1 expression via its canonical role in chromatin remodeling. Here, we report that TRIM28 through its B box domain increases L1 retrotransposition and facilitates shorter cDNA and L1 insert generation in cultured cells. Consistent with the latter, we observe that tumor specific L1 inserts are shorter in endometrial, ovarian, and prostate tumors with higher TRIM28 mRNA expression than in those with lower TRIM28 expression. We determine that three amino acids in the B box domain that are involved in TRIM28 multimerization are critical for its effect on both L1 retrotransposition and cDNA synthesis. We provideevidence that B boxes from the other two members in the Class VI TRIM proteins, TRIM24 and TRIM33, also increase L1 retrotransposition. Our findings could lead to a better understanding of the host/L1 evolutionary arms race in the germline and their interplay during tumorigenesis.

A Dual-Function "TRE-Lox" System for Genetic Deletion or Reversible, Titratable, and Near-Complete Downregulation of Cathepsin D.

Commonly employed methods for reversibly disrupting gene expression, such as those based on RNAi or CRISPRi, are rarely capable of achieving >80-90% downregulation, making them unsuitable for targeting genes that require more complete disruption to elicit a phenotype. Genetic deletion, on the other hand, while enabling complete disruption of target genes, often produces undesirable irreversible consequences such as cytotoxicity or cell death. Here we describe the design, development, and detailed characterization of a dual-function "TRE-Lox" system for effecting either (a) doxycycline (Dox)-mediated downregulation or (b) genetic deletion of a target gene-the lysosomal aspartyl protease cathepsin D (CatD)-based on targeted insertion of a tetracycline-response element (TRE) and two LoxP sites into the 5' end of the endogenous CatD gene (CTSD). Using an optimized reverse-tetracycline transrepressor (rtTR) variant fused with the Krüppel-associated box (KRAB) domain, we show that CatD expression can be disrupted by as much as 98% in mouse embryonic fibroblasts (MEFs). This system is highly sensitive to Dox (IC50 = 1.46 ng/mL) and results in rapid (t1/2 = 0.57 d) and titratable downregulation of CatD. Notably, even near-total disruption of CatD expression was completely reversed by withdrawal of Dox. As expected, transient expression of Cre recombinase results in complete deletion of the CTSD gene. The dual functionality of this novel system will facilitate future studies of the involvement of CatD in various diseases, particularly those attributable to partial loss of CatD function. In addition, the TRE-Lox approach should be applicable to the regulation of other target genes requiring more complete disruption than can be achieved by traditional methods.

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.

Abstract 1582: Inhibition of autophagy by CAGE targeting peptide retard tumor growth in non-small cell lung cancer of EGFR TKI resistance

Abstract Epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI) is the standard of care as 1st-line treatment for NSCLC patients with activating EGFR mutations (exon 19 deletion or L858R mutation). However, EGFR-TKI therapy eventually leads to acquired drug resistance approximately 1-2 years from the beginning of treatment. Multiple acquired resistance mechanisms including on-target EGFR mutations, phenotype transformation, and activation of bypass signaling molecules have been identified, but over 35% still remains unknown. CAGE, a cancer/testis antigen, was originally isolated from the sera of patients with gastric cancer and is known to regulate the autophagic flux in acquired resistance to EGFR TKI in NSCLC. Herein, the expression of CAGE was found in more than 50% of NSCLC patients using immunohistochemistry (IHC). Interestingly, the expression level was increased especially for the patients acquiring resistance to EGFR-TKIs. In addition, both PC-9/ER (Erlotinib-resistance cells) and H1975/OR (Osimertinib-resistant cells) cells demonstrated increased CAGE expression and autophagic flux, when compared with their parental cells. Thus, we developed the peptide drugs targeting the DEAD box domain of CAGE. This novel peptide effectively inhibited tumor cell proliferation in EGFR-TKI resistant cell line. Moreover, in Erlotinib-resistant patient-derived xenograft (PDX) mice, the peptide drug retarded tumor growth and enhanced anti-tumor efficacy. Collectively, our data suggest that CAGE plays a crucial role in the tumor progression and acquired resistance of EGFR TKI by the modulation of autophagic flux. Furthermore, we provided evidence that the inhibition of autophagy through CAGE- derived peptide could overcome the acquired resistance to EGFR TKIs in advanced/metastatic NSCLC patients. Citation Format: Jung-Young Shin, Min-Young Kim, Mi-Ran Lee, Hankyu Lee, Doyong Jeon, Seung Ryel Baek, Kim Joo Il, Jaemoon Koh, Dooil Jeoung, Jin Hyoung Kang. Inhibition of autophagy by CAGE targeting peptide retard tumor growth in non-small cell lung cancer of EGFR TKI resistance [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 1582.

Genome-wide identification, bioinformatics and expression analysis of HD-Zip gene family in peach

BackgroundHD-Zips (Homeodomain-Leucine Zippers) are a class of plant-specific transcription factors that play multiple roles in plant growth and development. Although some functions of HD-Zip transcription factor have been reported in several plants, it has not been comprehensively studied in peach, especially during adventitious root formation of peach cuttings.ResultsIn this study, 23 HD-Zip genes distributed on 6 chromosomes were identified from the peach (Prunus persica) genome, and named PpHDZ01-23 according to their positions on the chromosomes. These 23 PpHDZ transcription factors all contained a homeomorphism box domain and a leucine zipper domain, were divided into 4 subfamilies(I-IV) according to the evolutionary analysis, and their promoters contained many different cis-acting elements. Spatio-temporal expression pattern showed that these genes were expressed in many tissues with different levels, and they had distinct expression pattern during adventitious root formation and development.ConclusionOur results showed the roles of PpHDZs on root formation, which is helpful to better understand the classification and function of peach HD-Zip genes.

Molecular characterization and immune response of suppressor of cytokine signaling 5b from redlip mullet (Planiliza haematocheilus): Disclosing its anti-viral potential and effect on cell proliferation

The suppressor of cytokine signaling (SOCS) proteins family comprising eight proteins (SOCS1-7 and cytokine-inducible SH2-containing (CIS)) are classical feedback inhibitors of cytokine signaling. Although the biological role of CIS and SOCS1-3 have been extensively studied, the biological functions of SOCS4-7 remain unclear. Here, we elucidated the molecular characteristics, expression profile, immune response, anti-viral potential, and effect on cell proliferation of Phsocs5b, a member of the SOCS protein family from redlip mullet (Planiliza haematocheilus); phsocs5b comprised 1695 nucleotides. It was 564 amino acids long with a molecular weight of 62.3 kDa and a theoretical isoelectric point of 8.95. Like SOCS4-7 proteins, Phsocs5b comprised an SH2 domain, SOCS box domain, and a long N-terminal. SH2 domain is highly identical to its orthologs in other vertebrates. Phsocs5b, highly expressed in the brain tissue, was localized in the cytoplasm. Temporal changes in phsocs5b expression were observed following immune stimulation with polyinosinic: polycytidylic acid, lipopolysaccharide, and Lactococcus garvieae. In FHM cells, Phsocs5b overexpression suppressed the viral hemorrhagic septicemia virus (VHSV) infection and epidermal growth factor receptor (egfr) expression but increased the mRNA levels of pi3k, akt, pro-inflammatory cytokines (il1β and il8), and anti-viral genes (isg15 and ifn). Overall, our findings suggest that Phsocs5b attenuates VHSV infection, either by hindering the cell entry via degradation of Egfr, enhancing pro-inflammatory cytokines and anti-viral factor production, or both. The results also indicated that Phsocs5b could directly activate Pi3k/Akt pathway by itself, thus enhancing the proliferation and migration of cells. Taken together, Phsocs5b may be considered a potential therapeutic target to enhance immune responses while positively regulating the proliferation and migration of cells.

The Neurospora crassa PCL-1 cyclin is a PHO85-1 (PGOV) kinase partner that directs the complex to glycogen metabolism and is involved in calcium metabolism regulation.

Cyclins are a family of proteins characterized by possessing a cyclin box domain that mediates binding to cyclin dependent kinases (CDKs) partners. In this study, the search for a partner cyclin of the PHO85-1 CDK retrieved PCL-1 an ortholog of yeast Pcls (for Pho85 cyclins) that performs functions common to Pcls belonging to different cyclin families. We show here that PCL-1, as a typical cyclin, is involved in cell cycle control and cell progression. In addition, PCL-1 regulates glycogen metabolism; Δpcl-1 cells accumulate higher glycogen levels than wild-type cells and the glycogen synthase (GSN) enzyme is less phosphorylated and, therefore, more active in the mutant cells. Together with PHO85-1, PCL-1 phosphorylates in vitro GSN at the Ser636 amino acid residue. Modeling studies identified PHO85-1 and PCL-1 as a CDK/cyclin complex, with a conserved intermolecular region stabilized by hydrophobic and polar interactions. PCL-1 is also involved in calcium and NaCl stress response. Δpcl-1 cells are sensitive to high NaCl concentration; on the contrary, they grow better and overexpress calcium responsive genes under high calcium chloride concentration compared to the wild-type strain. The expression of the calcium-responsive CRZ-1 transcription factor is modulated by PCL-1, and this transcription factor seems to be less phosphorylated in Δpcl-1 cells since exhibits nuclear location in these cells in the absence of calcium. Our results show that PCL-1 locates at different cell regions suggesting that it may determine its activity by controlling its intracellular location and reveal an interesting functional divergence between yeast and filamentous fungus cyclins.

Zinc finger protein 251 deficiency impairs glucose metabolism by inducing adipocyte hypertrophy

Zinc finger protein (ZFP) 251 is a member of the C2H2 ZFP family containing a Krüppel-associated box domain that might mainly act as a transcriptional repressor. However, its cellular function remains largely unknown. Here, we discovered that ZFP251 deficiency caused glucose intolerance in mice. This phenotype was associated with impaired insulin signaling due to hypertrophic changes in white adipose tissue (WAT). Gene ontology analysis revealed that ZFP251 deficiency affected the expression of genes associated with adipocyte differentiation and lipid and fatty acid metabolism. Consistent with in vivo results, hypertrophic changes were observed in Zfp251 knockdown (KD) 3T3-L1 adipocytes. In addition, Zfp251 KD 3T3-L1 preadipocytes exhibited cell cycle arrest in G0/G1 phase, leading to impaired differentiation into mature adipocytes, upon which abnormal mitotic clonal expansion and reduced expression of adipogenic markers were exhibited. These results suggest that ZFP251 deficiency causes impaired adipogenesis and adipocyte hypertrophy, leading to dysfunction of WAT.

Spinocerebellar ataxia type 17-digenic TBP/STUB1 disease: neuropathologic features of an autopsied patient

Spinocerebellar ataxia (SCA) type 17-digenic TBP/STUB1 disease (SCA17-DI) has been recently segregated from SCA17, caused by digenic inheritance of two gene mutations – intermediate polyglutamine-encoding CAG/CAA repeat expansions (polyQ) in TBP (TBP41 − 49) and STUB1 heterozygosity – the former being associated with SCA17, and the latter with SCA48 and SCAR16 (autosomal recessive). In SCA17, most patients carry intermediate TBP41 − 49 alleles but show incomplete penetrance, and the missing heritability can be explained by a new entity whereby TBP41 − 49 requires the STUB1 variant to be symptomatic. The STUB1 gene encodes the chaperone-associated E3 ubiquitin ligase (CHIP) involved in ubiquitin-mediated proteasomal control of protein homeostasis. However, reports of the neuropathology are limited and role of STUB1 mutations in SCA17-DI remain unknown. Here we report the clinicopathologic features of identical twin siblings, one of whom was autopsied and was found to carry an intermediate allele (41 and 38 CAG/CAA repeats) in TBP and a heterozygous missense mutation in STUB1 (p.P243L). These patients developed autosomal recessive Huntington’s disease-like symptoms. Brain MRI showed diffuse atrophy of the cerebellum and T2WI revealed hyperintense lesions in the basal ganglia and periventricular deep white matter. The brain histopathology of the patient shared features characteristic of SCA17, such as degeneration of the cerebellar cortex and caudate nucleus, and presence of 1C2-positive neurons. Here we show that mutant CHIP fails to generate the polyubiquitin chain due to disrupted folding of the entire U box domain, thereby affecting the E3 activity of CHIP. When encountering patients with cerebellar ataxia, especially those with Huntington’s disease-like symptoms, genetic testing for STUB1 as well as TBP should be conducted for diagnosis of SCA17-DI, even in cases of sporadic or autosomal recessive inheritance.

KRAB family is involved in network shifts in response to osmotic stress in camels

ABSTRACT A feature of the camel is its tolerance to osmotic stress. However, few studies of osmotic stress in vivo or comparative analyses between different tissues of the camel have been performed. Here, we report the roles of Krüppel-associated box domain containing zinc-finger repressor proteins (KRAB-ZFPs) in transcriptional networks under osmotic stress in camels by analyzing transcriptomes of four different tissues under various osmotic conditions. We found that 273 of 278 KRAB-ZFPs were expressed in our data set, being involved in all of the 65 identified networks and exhibiting their extensive functional diversity. We also found that 110 KRAB-ZFPs were hub genes involved in more than half of the networks. We demonstrated that the osmotic stress response is involved in network shifts and that KRAB-ZFPs mediate this process. Finally, we presented the diverse mechanisms of osmotic stress responses in different tissues. These results revealed the genetic architecture of systematic physiological response in vivo to osmotic stress in camels. Our work will lead to new directions for studying the mechanism of osmotic stress response in anti-arid mammals.

AR-V7 exhibits non-canonical mechanisms of nuclear import and chromatin engagement in castrate-resistant prostate cancer.

Expression of the AR splice variant, androgen receptor variant 7 (AR-V7), in prostate cancer is correlated with poor patient survival and resistance to AR targeted therapies and taxanes. Currently, there is no specific inhibitor of AR-V7, while the molecular mechanisms regulating its biological function are not well elucidated. Here, we report that AR-V7 has unique biological features that functionally differentiate it from canonical AR-fl or from the second most prevalent variant, AR-v567. First, AR-V7 exhibits fast nuclear import kinetics via a pathway distinct from the nuclear localization signal dependent importin-α/β pathway used by AR-fl and AR-v567. We also show that the dimerization box domain, known to mediate AR dimerization and transactivation, is required for AR-V7 nuclear import but not for AR-fl. Once in the nucleus, AR-V7 is transcriptionally active, yet exhibits unusually high intranuclear mobility and transient chromatin interactions, unlike the stable chromatin association of liganded AR-fl. The high intranuclear mobility of AR-V7 together with its high transcriptional output, suggest a Hit-and-Run mode of transcription. Our findings reveal unique mechanisms regulating AR-V7 activity, offering the opportunity to develop selective therapeutic interventions.