Retinal Dehydrogenase Research Articles

Moderate Aerobic Exercise Induces Homeostatic IgA Generation in Senile Mice.

A T-cell-independent (TI) pathway activated by microbiota results in the generation of low-affinity homeostatic IgA with a critical role in intestinal homeostasis. Moderate aerobic exercise (MAE) provides a beneficial impact on intestinal immunity, but the action of MAE on TI-IgA generation under senescence conditions is unknown. This study aimed to determine the effects of long-term MAE on TI-IgA production in young (3 month old) BALB/c mice exercised until adulthood (6 months) or aging (24 months). Lamina propria (LP) from the small intestine was obtained to determine B cell and plasma cell sub-populations by flow cytometry and molecular factors related to class switch recombination [Thymic Stromal Lymphopoietin (TSLP), A Proliferation-Inducing Ligand (APRIL), B Cell Activating Factor (BAFF), inducible nitric oxide synthase (iNOS), and retinal dehydrogenase (RDH)] and the synthesis of IgA [α-chain, interleukin (IL)-6, IL-21, and Growth Factor-β (TGF-β)]; and epithelial cells evaluated IgA transitosis [polymeric immunoglobulin receptor (pIgR), tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), IL-4] by the RT-qPCR technique. The results were compared with data obtained from sedentary age-matched mice. Statistical analysis was computed with ANOVA, and p < 0.05 was considered to be a statistically significant difference. Under senescence conditions, MAE promoted the B cell and IgA+ B cells and APRIL, which may improve the intestinal response and ameliorate the inflammatory environment associated presumably with the downmodulation of pro-inflammatory mediators involved in the upmodulation of pIgR expression. Data suggested that MAE improved IgA and downmodulate the cytokine pro-inflammatory expression favoring homeostatic conditions in aging.

P141 Engineered probiotics treat inflammatory bowel disease by releasing all trans retinoic acid in the intestine

Abstract Background All trans retinoic acid (ATRA) is a potential candidate drug for the treatment of inflammatory bowel disease (IBD). ATRA can affect multiple pathways involved in the pathogenesis of IBD by binding to retinoic acid receptors. However, the shortcomings of poor compliance and significant side effects in patients who directly use ATRA limit its clinical application. Methods Here, we have developed a live biotherapeutic products (ELBPRA) based on synthetic biology methods for controllable delivery of ATRA on the gut. We first expressed four enzyme genes required for the synthesis of β-carotene, a precursor of retinoic acid, in Escherichia coli Nissle 1917 (EcN). In addition, we introduced the exogenous synthesis pathway of isopentane pyrophosphate into EcN to increase the yield of β-carotene. Then, β-carotene oxygenase and retinal dehydrogenase, were expressed in EcN to convert β-carotene into retinoic acid. However, considering the poor solubility of retinoic acid, we knocked out some genes related to vesicle formation in the EcN genome to increase extracellular production of retinoic acid. Finally, we investigated the therapeutic effect of ELBPRA using a colitis model induced by sodium dextran sulfate (DSS). Results ELBPRA continuously produces retinoic acid and delivers it in the form of vesicles. ELBPRA reduced weight loss and disease activity index in DSS-induced colitis. Similarly, the spleen index also demonstrates the effectiveness of ELBPRA in treating colitis. The ELBPRA intervention group reduced the degree of colon shortening in mice. In addition, ELBPRA improved crypt loss and disorder of crypt structure. Finally, we also evaluated the inflammatory cytokines in the serum. ELBPRA significantly increased the concentration of anti-inflammatory cytokine IL-10 and decreased the concentration of pro-inflammatory cytokine IL-1β and TNF-α. Conclusion Engineered probiotics ELBPRA can deliver retinoic acid in the form of vesicles to alleviate DSS-induced colitis in mice, which provides a safe and effective treatment strategy for ulcerative colitis.

Identification of approved drugs with ALDH1A1 inhibitory potential aimed at enhancing chemotherapy sensitivity in cancer cells: an in-silico drug repurposing approach

The aldehyde dehydrogenase 1A1 (ALDH1A1) also known as retinal dehydrogenase, is an enzyme normally involved in the cellular metabolism, development and detoxification processes in healthy cells. However, it’s also considered a cancer stem cell marker and its high levels of expression in several cancers, including breast, lung, ovarian, and colon cancer have been associated with poor prognosis and resistance to chemotherapy. Given its crucial role in chemotherapy resistance by detoxification of chemotherapeutic drugs, ALDH1A1 has attracted significant research interest as a potential therapeutic target for cancer. Though a few synthetic inhibitors of ALDH1A1 have been synthesized and their efficacy has been proved in-vitro and in-vivo studies, none of them have passed clinical trials so far. In this scenario, we have performed an in-silico study to verify whether any of the already approved drugs used for various purposes has the ability to inhibit catalytic activity of ALDH1A1, so that they can be repurposed for cancer therapy. Keeping in mind the feasibility of repurposing in a larger population we have selected the approved drugs from five widely used drug categories such as antibiotic, antiviral, antifungal, anti diabetic and antihypertensive for screening. Computational techniques like molecular docking, molecular dynamics simulations and MM-PBSA binding energy calculation have been used in this study to screen the approved drugs. Based on the logical analysis of results, we propose that three drugs – telmisartan, irbesartan and maraviroc can inhibit the catalytic activity of ALDH1A1 and thus can be repurposed to increase chemotherapy sensitivity in cancer cells. Communicated by Ramaswamy H. Sarma

Loss of Atg5 in Sertoli cells enhances the susceptibility of cadmium-impaired testicular spermatogenesis in mice.

Cadmium (Cd), one of the most common contaminants in diet and drinking water, impairs testicular germ cell development and spermatogenesis. Autophagy is essential for maintaining Sertoli cell function and Sertoli-germ cell communication. However, the role of Sertoli cell autophagy in Cd-caused spermatogenesis disorder remains unclear. Here, the mice of autophagy-related gene 5 (Atg5) knockouts in Sertoli cells were used to investigate the effect of autophagy deficiency on Cd-impaired spermatogenesis and its underlying mechanisms. Results showed that Sertoli cell-specific knockout of Atg5 exacerbated Cd-reduced sperm count and MVH (a specific marker for testicular germ cells) level in mice. Additionally, Sertoli cell Atg5 deficiency reduced the number of spermatocytes and decreased the level of meiosis-related proteins (SYCP3 and STRA8) in Cd-treated mouse testes. Loss of Atg5 in Sertoli cell exacerbated Cd-reduced the level of retinoic acid (RA) and retinal dehydrogenase (ALDH1A1 and ALDH1A) in mouse testes. Meanwhile, we found that the level of transcription factor WT1 was significantly downregulated in Atg5-/- plus Cd-treated testes. Further experiments showed that Wt1 overexpression restored Cd-decreased the levels of ALDH1A1 in Sertoli cells. Collectively, the above data suggest that knockout of Atg5 in Sertoli cell enhances the susceptibility of Cd-impaired testicular spermatogenesis. These findings provide new insights into autophagy of Sertoli cell preventing environmental toxicants-impaired testicular spermatogenesis.

Maximal clique centrality and bottleneck genes as novel biomarkers in ovarian cancer

ABSTRACT Ovarian cancer (OC) is second most common form of gynaecological cancer world wide . In this study, we collected and analyzed three ovarian cancer microarray raw datasets from Gene Expression Omnibus, NCBI, and identified a total of 1806 significant DEGs (Differentially expressed genes). The functional analysis of the DEGs showed that the 885 upregulated DEGs were mostly enriched in protein-binding activity, while the downregulated 796 genes were mostly enriched in retinal dehydrogenase activity and GABA receptor binding. We then constructed a protein–protein interaction network of the DEGs DEGs in ovarian cancer datasetsand analyzed the network to find cluster subnets, using molecular complex detection (MCODE). Common genes among top hub gene list, bottleneck gene list and maximum clique centrality (MCC) gene lists were identified as key driver genes, After analyzing the network. The following genes, STK12 (Serine threonine protein kinase), UBE2C (Ubiquitin-conjugating enzyme E2 C), CENPA (Centromere protein A), CCNB1 (Cyclin B1), POLD1 (polymerase delta 1) and KIF11 (Kinesin Family Member 11) were finally identified as driver genes. Higher expression of the key driver genes, STK12, UBE2C, CENPA, CCNB1, POLD1 and KIF11, was associated with lower overall survival (OS) among ovarian cancer patients. Therefore, the identified driver genes could be important diagnostic and prognostic biomarkers for predicting ovarian cancer progression and understanding the mechanism of tumour formation and recurrence.

Production of retinoic acid by engineered Saccharomyces cerevisiae using an endogenous aldehyde dehydrogenase

Retinoic acid (RA), a vitamin A (retinol)-derived lipophilic compound, is involved in various physiological functions. The demand for RA is growing in the pharmaceutical industry, but RA biosynthesis is still in its infancy compared to other forms of retinoids such as retinol and retinal, largely due to the lack of efficient retinal dehydrogenases. To achieve effective biosynthesis of RA, the catalytic activities of exogenous retinal dehydrogenases were comparatively analyzed in a previously constructed retinoids-producing Saccharomyces cerevisiae strain, followed by mining of endogenous enzymes with higher retinal dehydrogenase activities using homology-based search. After confirming the retinal oxidation activity of the endogenous aldehyde dehydrogenase Hfd1 using in vivo and in vitro experiments, it was overexpressed in multiple copies, and the resulting strain produced 99.71 mg/L of RA in shake-flask cultures. Finally, 545.28 mg/L of RA was produced in fed-batch fermentation. This study suggests the yeast endogenous Hfd1 as a potent catalyst for RA biosynthesis, and demonstrates the potential of yeast as a platform for RA production.

Proteomic Analysis of Endometrial Cancer Tissues from Patients with Type 2 Diabetes Mellitus.

Endometrial cancer (EC) is the most common form of gynecological cancer. Type 2 diabetes mellitus is associated with an increased risk of EC. Currently, no proteomic studies have investigated the role of diabetes in endometrial cancers from clinical samples. The present study aims to elucidate the molecular link between diabetes and EC using a proteomic approach. Endometrial tissue samples were obtained from age-matched patients (EC Diabetic and EC Non-Diabetic) during surgery. Untargeted proteomic analysis of the endometrial tissues was carried out using a two-dimensional difference in gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF). A total of 53 proteins were identified, with a significant difference in abundance (analysis of variance (ANOVA) test, p ≤ 0.05; fold-change ≥ 1.5) between the two groups, among which 30 were upregulated and 23 downregulated in the EC Diabetic group compared to EC Non-Diabetic. The significantly upregulated proteins included peroxiredoxin-1, vinculin, endoplasmin, annexin A5, calreticulin, and serotransferrin. The significantly downregulated proteins were myosin regulatory light polypeptide 9, Retinol dehydrogenase 12, protein WWC3, intraflagellar transport protein 88 homolog, superoxide dismutase [Cu-Zn], and retinal dehydrogenase 1. The network pathway was related to connective tissue disorder, developmental disorder, and hereditary disorder, with the identified proteins centered around dysregulation of ERK1/2 and F Actin signaling pathways. Cancer-associated protein alterations such as upregulation of peroxiredoxin-1, annexin 5, and iNOS, and downregulation of RDH12, retinaldehyde dehydrogenase 1, SOD1, and MYL 9, were found in the EC tissues of the diabetic group. Differential expression of proteins linked to cancer metastasis, such as the upregulation of vinculin and endoplasmin and downregulation of WWC3 and IFT88, was seen in the patients with diabetes. Calreticulin and alpha-enolase, which might have a role in the interplay between diabetes and EC, need further investigation.

The multiciliated cells in Rathke's cleft express CYP26A1 and respond to retinoic acid in the pituitary.

The adenohypophysis consists of the anterior and intermediate lobes (AL and IL). The marginal cell layer (MCL), including the ventral region of the IL and the dorsal region of the AL lining the Rathke's cleft, acts as the primary stem/progenitor cell niches in adult adenohypophysis. The cells of the MCL on the IL side consisted of cluster of differentiation 9 (CD9)-positive stem/progenitor cells with or without motile cilia. However, any additional cellular properties of multiciliated CD9-positive cells are not known. The present study aimed to identify the character of the multiciliated cells in stem cell niche of the pituitary gland. We observed the fine structure of the multiciliated cells in the MCL of male Wistar rats at an early stage after birth and in adulthood (P60) using scanning electron microscopy. Since the previous study showed that the MCL cells of adult rats synthesize retinoic acid (RA), the present study determined whether the multiciliated cells are involved in RA regulation by the expression of retinal aldehyde dehydrogenase 1 (RALDH1) and CYP26A1, an enzyme synthesizing and degrading RA, respectively. Results showed that 96% of multiciliated cells in adult male rats expressed CYP26A1, while 60% expressed RALDH1. Furthermore, the isolated CD9-positive cells from the IL side MCL responded to RA and activated the degradation system of RA by increasing Cyp26a1 expression. These findings indicated that multiciliated cells are involved in RA metabolism in the MCL. Our observations provide novel insights regarding the stem cell niche of the adult pituitary.

Syzygium aromaticum bud (clove) essential oil is a novel and safe aldose reductase inhibitor: in silico, in vitro, and in vivo evidence.

This study aimed to evaluate the antioxidant and antidiabetic properties of clove essential oil (CEO) and to elucidate its mode of action, using selected biochemical targets, relevant to diabetes, and, specifically, its inhibitory effect on the polyol pathway. In the current study, CEO was examined for its inhibitory effects on aldose reductase in silico, in vitro, and in vivo, as well as its antioxidative activity. In silico docking studies showed that all the selected major compounds of CEO have an energy change ranging between - 5.5 and - 8.8kcal/mol and an inhibition constant ranging between 357.08nM and 93.12µM. CEO significantly inhibits aldose reductase with an IC50 value of 58.55 ± 5.84µg/mL in a noncompetitive manner. The supplementation of CEO at 20mg/kg BW decreases retinal sorbitol dehydrogenase activity via decreased aldose reductase activity in streptozotocin (STZ)-induced diabetic Sprague Dawley rats. Moreover, diabetic rats injected with CEO have exhibited improved levels of glycemia. The IC50 values for ABTS, hydroxyl, and hydrogen peroxide scavenging activities of CEO were found to be 34.42, 277.4, and 39.99µg/mL, respectively. Reducing power assay and phosphomolybdate assay exhibited a reduction force with the A0.5 values of 50.25 and 140.16µg/mL, respectively. CEO potentially exerts a beneficial effect on diabetes-related complications due to its antioxidant and inhibitory effect on aldose reductase activity.

Novel variants in the RDH5 Gene in a Chinese Han family with fundus albipunctatus

BackgroundThe aim of this study is to identify the genetic defects in a Chinese family with fundus albipunctatus.MethodsComplete ophthalmic examinations, including slit-lamp biomicroscopy, dilated indirect ophthalmoscopy, fundus photography, autofluorescence, swept source optical coherence tomography (SS-OCT) and full-field electroretinography (ffERG) were performed. Genomic DNA was extracted from blood samples and whole genome sequencing was performed. Variants were validated with Sanger sequencing.ResultsSix members in this Chinese family, including three affected individuals and three controls, were recruited in this study. The ophthalmic examination of three recruited patients was consistent with fundus albipunctatus. Three variants, a novel frameshift deletion c.39delA [p.(Val14CysfsX47] and a haplotype of two rare missense variants, c.683G > A [p.(Arg228Gln)] along with c.710A > G [p.(Tyr237Cys], within the retinal dehydrogenase 5 (RDH5) gene were found to segregate with fundus albipunctatus in this family in an autosomal recessive matter.ConclusionWe identified novel compound heterozygous variants in RDH5 responsible for fundus albipunctatus in a large Chinese family. The results of our study further broaden the genetic defects of RDH5 associated with fundus albipunctatus.

Identification of eleven differentially expressed proteins in the tear film of patients with neovascular Age‐related Macular Degeneration (AMD)

AbstractPurposeThe exact mechanisms underlying the age‐related macular degeneration (AMD) are still not totally understood, but processes like inflammation, chronic oxidative stress (OS), and autophagy impairment, are currently believed to have strong influence of the disease development. There are multiple studies concerning changes of the proteome in different tissues in the course of AMD. As they all need invasive procedures to obtain a sample, we sought for a non‐invasive approach. Tear film can be a potential candidte for proteomic information regarding AMD.MethodsTear film was collected on Schirmer strips, 2D‐electrophoresis was performed, and Delta2D software was used for the identification of the differentially expressed proteins from the gels. The spots were excised, and identified by MALDI‐TOF mass spectrometer.Results30 patients were examined, 15 for each group. We have found 469 proteins in the analyzed samples, 11 differentiating proteins fulfilled the criteria of being statistically significant. 3 proteins were up‐regulated (Retinal dehydrogenase 1; ATP‐dependent translocase ABCB1; and alpha‐Enolase), and 8 were down‐regulated (Annexin A1; Annexin A4; Glutathione S‐transferase P; Aldo‐ketoreductase family 1 member A1; Allograft inflammatory factor 1; Cytospin‐A or Elongation factor 2; Short stature homeobox protein 2 and uncharacterized protein C11orf98).ConclusionsTear film can be used as a non‐invasively obtained source of proteomic information about cellular processes ongoing in the course of AMD. The differentially expressed proteins are involved in the choroidal neovascularization, inflammation, oxidative stress and autophagy impairment.

The rhodopsin-retinochrome system for retinal re-isomerization predates the origin of cephalopod eyes

BackgroundThe process of photoreception in most animals depends on the light induced isomerization of the chromophore retinal, bound to rhodopsin. To re-use retinal, the all-trans-retinal form needs to be re-isomerized to 11-cis-retinal, which can be achieved in different ways. In vertebrates, this mostly includes a stepwise enzymatic process called the visual cycle. The best studied re-isomerization system in protostomes is the rhodopsin-retinochrome system of cephalopods, which consists of rhodopsin, the photoisomerase retinochrome and the protein RALBP functioning as shuttle for retinal. In this study we investigate the expression of the rhodopsin-retinochrome system and functional components of the vertebrate visual cycle in a polyplacophoran mollusk, Leptochiton asellus, and examine the phylogenetic distribution of the individual components in other protostome animals.ResultsTree-based orthology assignments revealed that orthologs of the cephalopod retinochrome and RALBP are present in mollusks outside of cephalopods. By mining our dataset for vertebrate visual cycle components, we also found orthologs of the retinoid binding protein RLBP1, in polyplacophoran mollusks, cephalopods and a phoronid. In situ hybridization and antibody staining revealed that L. asellus retinochrome is co-expressed in the larval chiton photoreceptor cells (PRCs) with the visual rhodopsin, RALBP and RLBP1. In addition, multiple retinal dehydrogenases are expressed in the PRCs, which might also contribute to the rhodopsin-retinochrome system.ConclusionsWe conclude that the rhodopsin-retinochrome system is a common feature of mollusk PRCs and predates the origin of cephalopod eyes. Our results show that this system has to be extended by adding further components, which surprisingly, are shared with vertebrates.

Similarities in DSG1 and KRT3 Downregulation through Retinoic Acid Treatment and PAX6 Knockdown Related Expression Profiles: Does PAX6 Affect RA Signaling in Limbal Epithelial Cells?

Congenital PAX6-aniridia is a rare panocular disease resulting from limbal stem cell deficiency. In PAX6-aniridia, the downregulation of the retinol-metabolizing enzymes ADH7 (All-trans-retinol dehydrogenase 7) and ALDH1A1/A3 (Retinal dehydrogenase 1, Aldehyde dehydrogenase family 1 member A3) have been described in limbal epithelial cells (LECs) and conjunctival epithelial cells. The aim of this study was to identify the role of retinol derivates in the differentiation of human LEC and its potential impact on aniridia-associated keratopathy development. Human LEC were isolated from healthy donor corneas and were cultured with retinol, retinoic acid, or pan-retinoic acid receptor antagonist (AGN 193109) acting on RARα, β, γ (NR1B1, NR1B2 NR1B3) or were cultured with pan-retinoid X receptor antagonist (UVI 3003) acting on RXR α, β, γ (retinoid X receptor, NR2B1, NR2B2, BR2B3). Using qPCR, differentiation marker and retinoid-/fatty acid metabolism-related mRNA expression was analysed. DSG1 (Desmoglein 1), KRT3 (Keratin 3), and SPINK7 (Serine Peptidase Inhibitor Kazal Type 7) mRNA expression was downregulated when retinoid derivates were used. AGN 193109 treatment led to the upregulation of ADH7, KRT3, and DSG1 mRNA expression and to the downregulation of KRT12 (Keratin 12) and KRT19 (Keratin 19) mRNA expression. Retinol and all-trans retinoic acid affect some transcripts of corneal LEC in a similar way to what has been observed in the LEC of PAX6-aniridia patients with the altered expression of differentiation markers. An elevated concentration of retinol derivatives in LEC or an altered response to retinoids may contribute to this pattern. These initial findings help to explain ocular surface epithelia differentiation disorders in PAX6-aniridia and should be investigated in patient cells or in cell models in the future in more detail.

Estrogen regulates the expression of retinoic acid synthesis enzymes and binding proteins in mouse skin

Topical 17-beta-estradiol (E2) regulates the hair cycle, hair shaft differentiation, and sebum production. Vitamin A also regulates sebum production. Vitamin A metabolism proteins localized to the pilosebaceous unit (PSU; hair follicle and sebaceous gland); and were regulated by E2 in other tissues. This study tests the hypothesis that E2 also regulates vitamin A metabolism in the PSU. First, aromatase and estrogen receptors localized to similar sites as retinoid metabolism proteins during mid-anagen. Next, female and male wax stripped C57BL/6J mice were topically treated with E2, the estrogen receptor antagonist ICI 182,780 (ICI), letrozole, E2 plus letrozole, or vehicle control (acetone) during mid-anagen. E2 or one of its inhibitors regulated most of the vitamin A metabolism genes and proteins examined in a sex-dependent manner. Most components were higher in females and reduced with ICI in females. ICI reductions occurred in the premedulla, sebaceous gland, and epidermis. Reduced E2 also reduced RA receptors in the sebaceous gland and bulge in females. However, reduced E2 increased the number of retinal dehydrogenase 2 positive hair follicle associated dermal dendritic cells in males. These results suggest that estrogen regulates vitamin A metabolism in the skin. Interactions between E2 and vitamin A have implications in acne treatment, hair loss, and skin immunity.

A Chemoproteomics Approach to Determine the Mechanism of Testicular Toxicity for the Bruton’s Tyrosine Kinase Inhibitor CC-292

During drug development, potential safety issues can occur at any time. Understanding the cause of a toxicity can help with deciding on how to advance the drug development program. Chemoproteomics provides a way to help understand the cause of a toxicity wherein the affected tissue is accessible and can be probed with a covalently binding compound that is analogous to the offending drug. In this case, N-(3-(5-fluoro-2-(4-(2-methoxyethoxy)phenylamino)pyrimidin-4-ylamino)phenyl)acrylamide (CC-292), a covalently binding Bruton's tyrosine kinase inhibitor, had produced testicular toxicity in rodents. Experiments were conducted using a CC-292 analog that could be chemically modified with biotin to probe rodent testes homogenates for potential binding sites that were subsequently recovered with avidin beads. These biotin-tagged proteins undergo trypsin digest on the avidin beads to yield peptides that are identified using mass spectrometry. Two proteins were identified from the testicular homogenates of both rats and mice, namely retinal dehydrogenase 1 (ALDH1A1) and retinal dehydrogenase 2 (ALDH1A2). Literature confirmed a link between inhibition of these enzymes and testicular toxicity. Subsequently, molecular modeling was used to demonstrate that CC-292 can be docked into both the nicotinamide adenine dinucleotide and retinal binding pockets of the analogous human ALDH1A2 enzyme. These data suggest that the off-target binding site for CC-292 on retinal dehydrogenase enzymes may provide a mechanistic explanation to the testicular toxicity observed in rodents and that there may be a potential concern for human male fertility. SIGNIFICANCE STATEMENT: Biotinylated covalently binding drug analogues are used to enrich bound proteins from tissue homogenates wherein toxicity was observed in rodents. Bound proteins were subsequently identified by mass spectroscopy. Competition of the analog binding with the parent inhibitor itself and three-dimensional molecular modeling were used to establish a likely link between the off-targets of CC-292, ALDH1A1, and ALDH1A2 with potential testicular toxicity.

Retinal IMP Dehydrogenase Resist GTP Feedback Inhibition by Filamentous Polymerization

Dynamic assembly of metabolic enzymes is an evolutionarily conserved mechanism for the physical organization of metabolism. IMP dehydrogenase (IMPDH), is a highly-conserved enzyme that catalyzes the first committed step in GTP synthesis and forms dynamic filamentous assemblies in vertebrate cells. Humans have two isoforms of the protein: IMPDH2 which is upregulated in dividing cells and IMPDH1 which is expressed at low levels as the housekeeping enzyme in most cells. IMPDH2 assembles into filaments to tune feedback inhibition by the downstream product GTP, however the consequences of IMPDH1 polymerization are unclear. This question is made particularly compelling because there are 9 mutations in IMPDH1 that lead to blindness in humans, 8 of which cluster around the nucleotide binding sites that mediate filament assembly in IMPDH2. Moreover, IMPDH1 is produced as 2 unique splice variants in the retina, and their influence on assembly and regulation remain unexplored. Here we show that IMPDH1 polymerization does not confer increased resistance to GTP-inhibition, unlike the previously-studied isoform IMPDH2. We present high-resolution cryo-electron microscopy (cryoEM) structures that show IMPDH1 forms a novel filament when bound to GTP, providing a structural explanation for this difference in isoforms. Using kinetic assays we demonstrate that IMPDH1 retinal splice variants are four times less sensitive to GTP-inhibition than canonical IMPDH1, an effect that is specific to the retinal protein being assembled into a filament. We present a high-resolution cryoEM structure of one of the IMPDH1 retinal variants bound to GTP that shows it forms a filament identical to IMPDH2, explaining why polymerization increases resistance to GTP inhibition. We anticipate these findings to provide a foundation for understanding why mutations in IMPDH1 lead to blindness in humans and a more robust investigation of splice variants of other metabolic enzymes that form dynamic assemblies.

The effects of all-trans retinoic acid on immune cells and its formulation design for vaccines.

As one of the most important metabolites of vitamin A, all-trans retinoic acid (RA) plays a crucial role in regulating immune responses. RA has been shown to promote the differentiation of naïve T and B cells and perform diverse functions in the presence of different cytokines. RA also induces gut tropic lymphocytes through upregulating the expression of chemokine (C-C motif) receptor 9 (CCR9) and α4β7 integrin. In addition, RA promotes the expression of the enzyme retinal dehydrogenase (RALDH) on dendritic cells, which in turn strengthens the ability to synthesize RA. Due to the insolubility of RA, proper formulation design can maximize its ability to improve immune responses for vaccines. Recent studies have developed some formulations co-loading RA and antigen, which can effectively imprint lymphocytes gut homing properties and induce intestine immune responses as well as systemic responses through parenteral administration, providing a promising direction for the protection against mucosal infections. Here, we review the mechanism and effects of RA on lymphocyte differentiation and gut homing, and recent progress of RA delivery systems to improve mucosal immune responses.

Dietary Vitamin A Impacts Refractory Telogen.

Hair follicles cycle through periods of growth (anagen), regression (catagen), rest (telogen), and release (exogen). Telogen is further divided into refractory and competent telogen based on expression of bone morphogenetic protein 4 (BMP4) and wingless-related MMTV integration site 7A (WNT7A). During refractory telogen hair follicle stem cells (HFSC) are inhibited. Retinoic acid synthesis proteins localized to the hair follicle and this localization pattern changed throughout the hair cycle. In addition, excess retinyl esters arrested hair follicles in telogen. The purpose of this study was to further define these hair cycle changes. BMP4 and WNT7A expression was also used to distinguish refractory from competent telogen in C57BL/6J mice fed different levels of retinyl esters from two previous studies. These two studies produced opposite results; and differed in the amount of retinyl esters the dams consumed and the age of the mice when the different diet began. There were a greater percentage of hair follicles in refractory telogen both when mice were bred on an unpurified diet containing copious levels of retinyl esters (study 1) and consumed excess levels of retinyl esters starting at 12 weeks of age, as well as when mice were bred on a purified diet containing adequate levels of retinyl esters (study 2) and remained on this diet at 6 weeks of age. WNT7A expression was consistent with these results. Next, the localization of vitamin A metabolism proteins in the two stages of telogen was examined. Keratin 6 (KRT6) and cellular retinoic acid binding protein 2 (CRABP2) localized almost exclusively to refractory telogen hair follicles in study 1. However, KRT6 and CRABP2 localized to both competent and refractory telogen hair follicles in mice fed adequate and high levels of retinyl esters in study 2. In mice bred and fed an unpurified diet retinol dehydrogenase SDR16C5, retinal dehydrogenase 2 (ALDH1A2), and cytochrome p450 26B1 (CYP26B1), enzymes and proteins involved in RA metabolism, localized to BMP4 positive refractory telogen hair follicles. This suggests that vitamin A may contribute to the inhibition of HFSC during refractory telogen in a dose dependent manner.

PRMT3 interacts with ALDH1A1 and regulates gene-expression by inhibiting retinoic acid signaling

Protein arginine methyltransferase 3 (PRMT3) regulates protein functions by introducing asymmetric dimethylation marks at the arginine residues in proteins. However, very little is known about the interaction partners of PRMT3 and their functional outcomes. Using yeast-two hybrid screening, we identified Retinal dehydrogenase 1 (ALDH1A1) as a potential interaction partner of PRMT3 and confirmed this interaction using different methods. ALDH1A1 regulates variety of cellular processes by catalyzing the conversion of retinaldehyde to retinoic acid. By molecular docking and site-directed mutagenesis, we identified the specific residues in the catalytic domain of PRMT3 that facilitate interaction with the C-terminal region of ALDH1A1. PRMT3 inhibits the enzymatic activity of ALDH1A1 and negatively regulates the expression of retinoic acid responsive genes in a methyltransferase activity independent manner. Our findings show that in addition to regulating protein functions by introducing methylation modifications, PRMT3 could also regulate global gene expression through protein-protein interactions.

COVID-19 disease and malignant cancers: The impact for the furin gene expression in susceptibility to SARS-CoV-2.

Furin is a proprotein convertase that activates different kinds of regulatory proteins, including SARS-CoV-2 spike protein which contains an additional furin-specific cleavage site. It is essential in predicting cancer patients' susceptibility to SARS-CoV-2 and the disease outcomes due to varying furin expressions in tumor tissues. In this study, we analyzed furin's expression, methylation, mutation rate, functional enrichment, survival rate and COVID-19 outcomes in normal and cancer tissues using online databases, and our IHC. As a result, furin presented with biased expression profiles in normal tissues, showing 12.25-fold higher than ACE2 in the lungs. The furin expression in tumors were significantly increased in ESCA and TGCT, and decreased in DLBC and THYM, indicating furin may play critical mechanistic functions in COVID-19 viral entry into cells in these cancer patients. Line with furin over/downexpression, furin promoter hypo-/hyper-methylation may be the regulatory cause of disease and lead to pathogenesis of ESCA and THYM. Furthermore, presence of FURIN-201 isoform with functional domains (P_proprotein, Peptidase_S8 and S8_pro-domain) is highest in all cancer types in comparison to other isoforms, demonstrating its use in tumorigenesis and SARS-Cov-2 entry into tumor tissues. Furin mutation frequency was highest in UCES, and its mutation might elevate ACE2 expression in LUAD and UCEC, reduce ACE2 expression in COAD, elevate HSPA5 expression in PAAD, and elevate TMPRSS2 expression in BRCA. These results showed that furin mutations mostly increased expression of ACE2, HSPA5, and TMPRSS2 in certain cancers, indicating furin mutations might facilitate COVID-19 cell entry in cancer patients. In addition, high expression of furin was significantly inversely correlated with long overall survival (OS) in LGG and correlated with long OS in COAD and KIRC, indicating that it could be used as a favorable prognostic marker for cancer patients' survival. GO and KEGG demonstrated that furin was mostly enriched in genes for metabolic and biosynthetic processes, retinal dehydrogenase activity, tRNA methyltransferase activity, and genes involving COVID-19, further supporting its role in COVID-19 and cancer metabolism. Moreover, Cordycepin (CD) inhibited furin expression in a dosage dependent manner. Altogether, furin's high expression might not only implies increased susceptibility to SARS-CoV-2 and higher severity of COVID-19 symptoms in cancer patients, but also it highlights the need for cancer treatment and therapy during the COVID-19 pandemic. CD might have a potential to develop an anti-SARS-CoV-2 drug through inhibiting furin expression.