Biochemical communication between filament-forming enzymes: Potential Regulatory Roles of Metabolites in Enzyme Co-assemblies with CTP Synthase.

A host of metabolic enzymes reversibly self-assemble to form membrane-less, intracellular filaments under normal physiological conditions and in response to stress. Often, these enzymes reside at metabolic control points, suggesting that filament formation affords an additional regulatory mechanism. Examples include cytidine-5'-triphosphate (CTP) synthase (CTPS), which catalyzes the rate-limiting step for the de novo biosynthesis of CTP; inosine-5'-monophosphate dehydrogenase (IMPDH), which controls biosynthetic access to guanosine-5'-triphosphate (GTP); and ∆1-pyrroline-5-carboxylate (P5C) synthase (P5CS) that catalyzes the formation of P5C, which links the Krebs cycle, urea cycle, and proline metabolism. Intriguingly, CTPS can exist in co-assemblies with IMPDH or P5CS. Since GTP is an allosteric activator of CTPS, the association of CTPS and IMPDH filaments accords with the need to coordinate pyrimidine and purine biosynthesis. Herein, a hypothesis is presented furnishing a biochemical connection underlying co-assembly of CTPS and P5CS filaments - potent inhibition of CTPS by glutamate γ-semialdehyde, the open-chain form of P5C.

IMPDH2 forms spots at branching sites and distal ends of astrocyte stem processes.

Inosine monophosphate dehydrogenase (IMPDH) is a rate-limiting enzyme in the de novo GTP biosynthesis pathway. Recent studies suggest that IMPDH2, an isoform of IMPDH, can localize to specific subcellular compartments under certain conditions and regulate site-specific GTP availability and small GTPase activity in invasive cancer cells. However, it is unclear whether IMPDH2 plays a site-specific regulatory role in subcellular functions in healthy cells. In this study, we focused on brain cells and examined the localization pattern of IMPDH2. We discovered that IMPDH2 forms localized spots in the astrocytes of the adult mouse hippocampus. Further analysis of spot distribution in primary astrocyte cultures revealed that IMPDH2 spots are predominantly localized on branching sites and distal ends of astrocyte stem processes. Our findings suggest a potential unidentified role for IMPDH2 and GTP synthesis specifically at specialized nodes of astrocyte branches.

CLOCK Acetylates ASS1 to Drive Circadian Rhythm of Ureagenesis.

In addition to responding to environmental entrainment with diurnal variation, metabolism is also tightly controlled by cell-autonomous circadian clock. Extensive studies have revealed key roles of transcription in circadian control. Post-transcriptional regulation for the rhythmic gating of metabolic enzymes remains elusive. Here, we show that arginine biosynthesis and subsequent ureagenesis are collectively regulated by CLOCK (circadian locomotor output cycles kaput) in circadian rhythms. Facilitated by BMAL1 (brain and muscle Arnt-like protein), CLOCK directly acetylates K165 and K176 of argininosuccinate synthase (ASS1) to inactivate ASS1, which catalyzes the rate-limiting step of arginine biosynthesis. ASS1 acetylation by CLOCK exhibits circadian oscillation in human cells and mouse liver, possibly caused by rhythmic interaction between CLOCK and ASS1, leading to the circadian regulation of ASS1 and ureagenesis. Furthermore, we also identified NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 9 (NDUFA9) and inosine-5'-monophosphate dehydrogenase 2 (IMPDH2) as acetylation substrates of CLOCK. Taken together, CLOCK modulates metabolic rhythmicity by acting as a rhythmic acetyl-transferase for metabolic enzymes.

The IMPDH cytoophidium couples metabolism and fetal development in mice.

The cytoophidium is an evolutionarily conserved subcellular structure formed by filamentous polymers of metabolic enzymes. In vertebrates, inosine monophosphate dehydrogenase (IMPDH), which catalyses the rate-limiting step in guanosine triphosphate (GTP) biosynthesis, is one of the best-known cytoophidium-forming enzymes. Formation of the cytoophidium has been proposed to alleviate the inhibition of IMPDH, thereby facilitating GTP production to support the rapid proliferation of certain cell types such as lymphocytes, cancer cells and pluripotent stem cells (PSCs). However, past studies lacked appropriate models to elucidate the significance of IMPDH cytoophidium under normal physiological conditions. In this study, we demonstrate that the presence of IMPDH cytoophidium in mouse PSCs correlates with their metabolic status rather than pluripotency. By introducing IMPDH2 Y12C point mutation through genome editing, we established mouse embryonic stem cell (ESC) lines incapable of forming IMPDH polymers and the cytoophidium. Our data indicate an important role of IMPDH cytoophidium in sustaining a positive feedback loop that couples nucleotide biosynthesis with upstream metabolic pathways. Additionally, we find that IMPDH2 Y12C mutation leads to decreased cell proliferation and increased DNA damage in teratomas, as well as impaired embryo development following blastocoel injection. Further analysis shows that IMPDH cytoophidium assembly in mouse embryonic development begins after implantation and gradually increases throughout fetal development. These findings provide insights into the regulation of IMPDH polymerisation in embryogenesis and its significance in coordinating cell metabolism and development.

Neurodevelopmental disorder mutations in the purine biosynthetic enzyme IMPDH2 disrupt its allosteric regulation.

Inosine 5' monophosphate dehydrogenase (IMPDH) is a critical regulatory enzyme in purine nucleotide biosynthesis that is inhibited by the downstream product GTP. Multiple point mutations in the human isoform IMPDH2 have recently been associated with dystonia and other neurodevelopmental disorders, but the effect of the mutations on enzyme function has not been described. Here, we report the identification of two additional missense variants in IMPDH2 from affected individuals and show that all of the disease-associated mutations disrupt GTP regulation. Cryo-EM structures of one IMPDH2 mutant suggest this regulatory defect arises from a shift in the conformational equilibrium toward a more active state. This structural and functional analysis provides insight into IMPDH2-associated disease mechanisms that point to potential therapeutic approaches and raises new questions about fundamental aspects of IMPDH regulation.

Deletion of Impdh2 in adipocyte precursors limits the expansion of white adipose tissue and enhances metabolic health with overnutrition.

The equilibrium between the hypertrophic growth of existing adipocytes and adipogenesis is vital in managing metabolic stability in white adipocytes when faced with overnutrition. Adipogenesis has been established as a key player in combating metabolic irregularities caused by various factors. However, the benefits of increasing adipogenesis-mediated white adipose tissue (WAT) expansion for metabolic health regulation remain uncertain. Our findings reveal an increase in Impdh2 expression during the adipogenesis phase, both in vivo and in vitro. Xmp enhances adipogenic potential by fostering mitotic clonal expansion (MCE). The conditional knockout of Impdh2 in adipocyte progenitor cells(APCs) in adult and aged mice effectively curbs white adipose tissue expansion, ameliorates glucose tolerance, and augments energy expenditure under high-fat diet (HFD). However, no significant difference is observed under normal chow diet (NCD). Concurrently, the knockout of Impdh2 in APCs significantly reduces the count of new adipocytes induced by HFD, without affecting adipocyte size. Mechanistically, Impdh2 regulates the proliferation of APCs during the MCE phase via Xmp. Exogenous Xmp can significantly offset the reduction in adipogenic abilities of APCs due to Impdh2 deficiency. In summary, we discovered that adipogenesis-mediated WAT expansion, induced by overnutrition, also contributes to metabolic abnormalities. Moreover, the pivotal role of Impdh2 in regulating adipogenesis in APCs offers a novel therapeutic approach to combat obesity.

Impdh2 deficiency suppresses osteoclastogenesis through mitochondrial oxidative phosphorylation and alleviates ovariectomy-induced osteoporosis.

Abnormalities in osteoclastic generation or activity disrupt bone homeostasis and are highly involved in many pathologic bone-related diseases, including rheumatoid arthritis, osteopetrosis, and osteoporosis. Control of osteoclast-mediated bone resorption is crucial for treating these bone diseases. However, the mechanisms of control of osteoclastogenesis are incompletely understood. In this study, we identified that inosine 5'-monophosphate dehydrogenase type II (Impdh2) positively regulates bone resorption. By histomorphometric analysis, Impdh2 deletion in mouse myeloid lineage cells (Impdh2LysM-/- mice) showed a high bone mass due to the reduced osteoclast number. qPCR and western blotting results demonstrated that the expression of osteoclast marker genes, including Nfatc1, Ctsk, Calcr, Acp5, Dcstamp, and Atp6v0d2, was significantly decreased in the Impdh2LysM-/- mice. Furthermore, the Impdh inhibitor MPA treatment inhibited osteoclast differentiation and induced Impdh2-cytoophidia formation. The ability of osteoclast differentiation was recovered after MPA deprivation. Interestingly, genome-wide analysis revealed that the osteoclastic mitochondrial biogenesis and functions, such as oxidative phosphorylation, were impaired in the Impdh2LysM-/- mice. Moreover, the deletion of Impdh2 alleviated ovariectomy-induced bone loss. In conclusion, our findings revealed a previously unrecognized function of Impdh2, suggesting that Impdh2-mediated mechanisms represent therapeutic targets for osteolytic diseases.

Inosine monophosphate dehydrogenase type 2 polymorphism IMPDH2 3757T>C (rs11706052) and 12-month evolution of the graft function in renal transplant recipients on mycophenolate-based immunosuppression.

Variant allele at the inosine monophosphate dehydrogenase type 2 polymorphism IMPDH2 3757T>C has been associated with increased enzyme activity and reduced susceptibility to mycophenolic acid (MPA) in vitro. It has been suggested associated with an increased risk of acute rejection in renal transplant recipients on MPA-based immunosuppression, but not unambiguously. We assessed one-year evolution of the estimated glomerular filtration rate (eGFR) in transplanted variant allele carriers and wild-type subjects, while controlling for a number of demographic, pharmacogenetic, (co)morbidity, and treatment baseline and time-varying covariates. The eGFR slopes to day 28 (GMR = 1.01, 95% CI 0.93-1.09), and between days 28 and 365 (GMR = 1.01, 95% CI 0.99-1.02) were practically identical in 52 variant carriers and 202 wild-type controls. The estimates (95%CIs) remained within the limits of ±20% difference even after adjustment for a strong hypothetical effect of unmeasured confounders. Polymorphism IMPDH2 3757T>C does not affect the renal graft function over the 1st year after transplantation.

Wnt/ß-catenin signalling activates IMPDH2-mediated purine metabolism to facilitate oxaliplatin resistance by inhibiting caspase-dependent apoptosis in colorectal cancer.

BACKGROUND: Oxaliplatin resistance usually leads to therapeutic failure and poor prognosis in colorectal cancer (CRC), while the underlying mechanisms are not yet fully understood. Metabolic reprogramming is strongly linked to drug resistance, however, the role and mechanism of metabolic reprogramming in oxaliplatin resistance remain unclear. Here, we aim to explore the functions and mechanisms of purine metabolism on the oxaliplatin-induced apoptosis of CRC. METHODS: An oxaliplatin-resistant CRC cell line was generated, and untargeted metabolomics analysis was conducted. The inosine 5'-monophosphate dehydrogenase type II (IMPDH2) expression in CRC cell lines was determined by quantitative real-time polymerase chain reaction (qPCR) and western blotting analysis. The effects of IMPDH2 overexpression, knockdown and pharmacological inhibition on oxaliplatin resistance in CRC were assessed by flow cytometry analysis of cell apoptosis in vivo and in vitro. RESULTS: Metabolic analysis revealed that the levels of purine metabolites, especially guanosine monophosphate (GMP), were markedly elevated in oxaliplatin-resistant CRC cells. The accumulation of purine metabolites mainly arose from the upregulation of IMPDH2 expression. Gene set enrichment analysis (GSEA) indicated high IMPDH2 expression in CRC correlates with PURINE_METABOLISM and MULTIPLE-DRUG-RESISTANCE pathways. CRC cells with higher IMPDH2 expression were more resistant to oxaliplatin-induced apoptosis. Overexpression of IMPDH2 in CRC cells resulted in reduced cell death upon treatment with oxaliplatin, whereas knockdown of IMPDH2 led to increased sensitivity to oxaliplatin through influencing the activation of the Caspase 7/8/9 and PARP1 proteins on cell apoptosis. Targeted inhibition of IMPDH2 by mycophenolic acid (MPA) or mycophenolate mofetil (MMF) enhanced cell apoptosis in vitro and decreased in vivo tumour burden when combined with oxaliplatin treatment. Mechanistically, the Wnt/ß-catenin signalling was hyperactivated in oxaliplatin-resistant CRC cells, and a reciprocal positive regulatory mechanism existed between Wnt/ß-catenin and IMPDH2. Blocking the Wnt/ß-catenin pathway could resensitize resistant cells to oxaliplatin, which could be restored by the addition of GMP. CONCLUSIONS: IMPDH2 is a predictive biomarker and therapeutic target for oxaliplatin resistance in CRC.

Clonal evolution mechanisms in NT5C2 mutant-relapsed acute lymphoblastic leukaemia.

Relapsed acute lymphoblastic leukaemia (ALL) is associated with resistance to chemotherapy and poor prognosis. Gain-of-function mutations in the 5'-nucleotidase, cytosolic II (NT5C2) gene induce resistance to 6-mercaptopurine and are selectively present in relapsed ALL. Yet, the mechanisms involved in NT5C2 mutation-driven clonal evolution during the initiation of leukaemia, disease progression and relapse remain unknown. Here we use a conditional-and-inducible leukaemia model to demonstrate that expression of NT5C2(R367Q), a highly prevalent relapsed-ALL NT5C2 mutation, induces resistance to chemotherapy with 6-mercaptopurine at the cost of impaired leukaemia cell growth and leukaemia-initiating cell activity. The loss-of-fitness phenotype of NT5C2+/R367Q mutant cells is associated with excess export of purines to the extracellular space and depletion of the intracellular purine-nucleotide pool. Consequently, blocking guanosine synthesis by inhibition of inosine-5'-monophosphate dehydrogenase (IMPDH) induced increased cytotoxicity against NT5C2-mutant leukaemia lymphoblasts. These results identify the fitness cost of NT5C2 mutation and resistance to chemotherapy as key evolutionary drivers that shape clonal evolution in relapsed ALL and support a role for IMPDH inhibition in the treatment of ALL.

Asymmetric inheritance of cytoophidia could contribute to determine cell fate and plasticity: The onset of alternative differentiation patterns in daughter cells may rely on the acquisition of either CTPS or IMPDH cytoophidia: The onset of alternative differentiation patterns in daughter cells may rely on the acquisition of either CTPS or IMPDH cytoophidia.

Two enzymes involved in the synthesis of pyrimidine and purine nucleotides, CTP synthase (CTPS) and IMP dehydrogenase (IMPDH), can assemble into a single or very few large filaments called rods and rings (RR) or cytoophidia. Most recently, asymmetric cytoplasmic distribution of organelles during cell division has been described as a decisive event in hematopoietic stem cell fate. We propose that cytoophidia, which could be considered as membrane-less organelles, may also be distributed asymmetrically during mammalian cell division as previously described for Schizosaccharomyces pombe. Furthermore, because each type of nucleotide intervenes in distinct processes (e.g., membrane synthesis, glycosylation, and G protein-signaling), alterations in the rate of synthesis of specific nucleotide types could influence cell differentiation in multiple ways. Therefore, we hypothesize that whether a daughter cell inherits or not CTPS or IMPDH filaments determines its fate and that this asymmetric inheritance, together with the dynamic nature of these structures enables plasticity in a cell population.

The interaction between the Spt6-tSH2 domain and Rpb1 affects multiple functions of RNA Polymerase II.

The conserved transcription elongation factor Spt6 makes several contacts with the RNA Polymerase II (RNAPII) complex, including a high-affinity interaction between the Spt6 tandem SH2 domain (Spt6-tSH2) and phosphorylated residues of the Rpb1 subunit in the linker between the catalytic core and the C-terminal domain (CTD) heptad repeats. This interaction contributes to generic localization of Spt6, but we show here that it also has gene-specific roles. Disrupting the interface affected transcription start site selection at a subset of genes whose expression is regulated by this choice, and this was accompanied by changes in a distinct pattern of Spt6 accumulation at these sites. Splicing efficiency was also diminished, as was apparent progression through introns that encode snoRNAs. Chromatin-mediated repression was impaired, and a distinct role in maintaining +1 nucleosomes was identified, especially at ribosomal protein genes. The Spt6-tSH2:Rpb1 interface therefore has both genome-wide functions and local roles at subsets of genes where dynamic decisions regarding initiation, transcript processing, or termination are made. We propose that the interaction modulates the availability or activity of the core elongation and histone chaperone functions of Spt6, contributing to coordination between RNAPII and its accessory factors as varying local conditions call for dynamic responses.

IMPDH Inhibition Decreases TERT Expression and Synergizes the Cytotoxic Effect of Chemotherapeutic Agents in Glioblastoma Cells.

IMP dehydrogenase (IMPDH) inhibition has emerged as a new target therapy for glioblastoma multiforme (GBM), which remains one of the most refractory tumors to date. TCGA analyses revealed distinct expression profiles of IMPDH isoenzymes in various subtypes of GBM and low-grade glioma (LGG). To dissect the mechanism(s) underlying the anti-tumor effect of IMPDH inhibition in adult GBM, we investigated how mycophenolic acid (MPA, an IMPDH inhibitor) treatment affected key oncogenic drivers in glioblastoma cells. Our results showed that MPA decreased the expression of telomerase reverse transcriptase (TERT) in both U87 and U251 cells, and the expression of O6-methylguanine-DNA methyltransferase (MGMT) in U251 cells. In support, MPA treatment reduced the amount of telomere repeats in U87 and U251 cells. TERT downregulation by MPA was associated with a significant decrease in c-Myc (a TERT transcription activator) in U87 but not U251 cells, and a dose-dependent increase in p53 and CCCTC-binding factor (CTCF) (TERT repressors) in both U87 and U251 cells. In U251 cells, MPA displayed strong cytotoxic synergy with BCNU and moderate synergy with irinotecan, oxaliplatin, paclitaxel, or temozolomide (TMZ). In U87 cells, MPA displayed strong cytotoxic synergy with all except TMZ, acting primarily through the apoptotic pathway. Our work expands the mechanistic potential of IMPDH inhibition to TERT/telomere regulation and reveals a synthetic lethality between MPA and anti-GBM drugs.

[Analysis of a patient with Retinitis pigmentosa due to a novel variant of IMPDH1 gene].

OBJECTIVE: To explore the genetic basis for a patient with autosomal dominant retinitis pigmentosa (RP). METHODS: A male patient with RP treated at Gansu Provincial Maternal and Child Health Care Hospital in September 2019 was selected as the study subject. Clinical data was collected. Peripheral blood samples of the patient and his parents were subjected to whole exome sequencing (WES). Candidate variant was validated by Sanger sequencing and bioinformatic analysis. RESULTS: The patient, a 29-year-old male, developed night blindness, amblyopia, visual field defects and optic disc abnormalities since childhood. Gene sequencing revealed that he has harbored a heterozygous c.942G>C (p.Lys314Asn) variant of the IMPDH1 gene, which was inherited from his mother, whilst his father was of the wild type. Based on the guidelines from the American College of Medical Genetics and Genomics, the c.942G>C variant was predicted as likely pathogenic (PM1+PM2_Supporting+PP3+PP1). CONCLUSION: The c.942G>C (p.Lys314Asn) variant in the IMPDH1 gene probably underlay the RP in this patient.

A highly conserved zebrafish IMPDH retinal isoform produces the majority of guanine and forms dynamic protein filaments in photoreceptor cells.

Inosine monophosphate dehydrogenase (IMPDH) is a key regulatory enzyme in the de novo synthesis of the purine base guanine. Dominant mutations in human IMPDH1 cause photoreceptor degeneration for reasons that are unknown. Here, we sought to provide some foundational information on Impdh1a in the zebrafish retina. We found that in zebrafish, gene subfunctionalization due to ancestral duplication resulted in a predominant retinal variant expressed exclusively in rod and cone photoreceptors. This variant is structurally and functionally similar to the human IMPDH1 retinal variant and shares a reduced sensitivity to GTP-mediated inhibition. We also demonstrated that Impdh1a forms prominent protein filaments in vitro and in vivo in both rod and cone photoreceptor cell bodies, synapses, and to a lesser degree, in outer segments. These filaments changed length and cellular distribution throughout the day consistent with diurnal changes in both mRNA and protein levels. The loss of Impdh1a resulted in a substantial reduction of guanine levels, although cellular morphology and cGMP levels remained normal. Our findings demonstrate a significant role for IMPDH1 in photoreceptor guanine production and provide fundamental new information on the details of this protein in the zebrafish retina.

Molecular crowding facilitates bundling of IMPDH polymers and cytoophidium formation.

The cytoophidium is a unique type of membraneless compartment comprising of filamentous protein polymers. Inosine monophosphate dehydrogenase (IMPDH) catalyzes the rate-limiting step of de novo GTP biosynthesis and plays critical roles in active cell metabolism. However, the molecular regulation of cytoophidium formation is poorly understood. Here we show that human IMPDH2 polymers bundle up to form cytoophidium-like aggregates in vitro when macromolecular crowders are present. The self-association of IMPDH polymers is suggested to rely on electrostatic interactions. In cells, the increase of molecular crowding with hyperosmotic medium induces cytoophidia, while the decrease of that by the inhibition of RNA synthesis perturbs cytoophidium assembly. In addition to IMPDH, CTPS and PRPS cytoophidium could be also induced by hyperosmolality, suggesting a universal phenomenon of cytoophidium-forming proteins. Finally, our results indicate that the cytoophidium can prolong the half-life of IMPDH, which is proposed to be one of conserved functions of this subcellular compartment.

Novel Variant IMPDH1 c.134A>G, p.(Tyr45Cys): Phenotype-Genotype Correlation Revealed Likely Benign Clinical Significance.

Pathogenic variants in IMPDH1 are associated with autosomal dominant retinitis pigmentosa 10 (RP10), and Leber congenital amaurosis 11. This case report of a 13-year-old girl with Down's syndrome and keratoglobus is aimed at linking the novel variant IMPDH1 c.134A>G, p.(Tyr45Cys), a variant of uncertain significance, to a clinical phenotype and to provide grounds for the objective assignment of its benign features. RP10 is characterized by the early onset and rapid progression of ocular symptoms, beginning with nyctalopia in childhood, accompanied by typical RP fundus changes. As evidenced via thorough clinical examination and testing, none of the RP10 characteristics were present in our patient. On the contrary, our patient who was heterozygous for IMPDH1 c.134A>G, p.(Tyr45Cys) showed no signs of peripheral retinal dystrophy, and did not manifest any disease characteristics typical of the IMPDH1 gene mutation. Consequently, we conclude that the variant did not contribute to the phenotype. According to standards and guidelines for the interpretation of sequence variants, IMPDH1 c.134A>G, p.(Tyr45Cys) revealed likely benign features.

IMPDH forms the cytoophidium in zebrafish.

Inosine monophosphate dehydrogenase (IMPDH) catalyzes the rate-limiting step in de novo guanine nucleotide biosynthesis. Its activity is negatively regulated by the binding of GTP. IMPDH can form a membraneless subcellular structure termed the cytoophidium in response to certain changes in the metabolic status of the cell. The polymeric form of IMPDH, which is the subunit of the cytoophidium, has been shown to be more resistant to the inhibition by GTP at physiological concentrations, implying a functional correlation between cytoophidium formation and the upregulation of GTP biosynthesis. Herein we demonstrate that zebrafish IMPDH1b and IMPDH2 isoforms can assemble abundant cytoophidium in most of cultured cells under stimuli, while zebrafish IMPDH1a shows distinctive properties of forming the cytoophidium in different cell types. Point mutations that disrupt cytoophidium structure in mammalian models also prevent the aggregation of zebrafish IMPDHs. In addition, we discover the presence of the IMPDH cytoophidium in various tissues of larval and adult fish under normal growth conditions. Our results reveal that polymerization and cytoophidium assembly of IMPDH can be a regulatory machinery conserved among vertebrates, and with specific physiological purposes.

IMPDH dysregulation in disease: a mini review.

Inosine-5'-monophosphate dehydrogenase (IMPDH) is a highly conserved enzyme in purine metabolism that is tightly regulated on multiple levels. IMPDH has a critical role in purine biosynthesis, where it regulates flux at the branch point between adenine and guanine nucleotide synthesis, but it also has a role in transcription regulation and other moonlighting functions have been described. Vertebrates have two isoforms, IMPDH1 and IMPDH2, and point mutations in each are linked to human disease. Mutations in IMPDH2 in humans are associated with neurodevelopmental disease, but the effects of mutations at the enzyme level have not yet been characterized. Mutations in IMPDH1 lead to retinal degeneration in humans, and recent studies have characterized how they cause functional defects in regulation. IMPDH1 is expressed as two unique splice variants in the retina, a tissue with very high and specific demands for purine nucleotides. Recent studies have revealed functional differences among splice variants, demonstrating that retinal variants up-regulate guanine nucleotide synthesis by reducing sensitivity to feedback inhibition by downstream products. A better understanding of the role of IMPDH1 in the retina and the characterization of an animal disease model will be critical for determining the molecular mechanism of IMPDH1-associated blindness.

MYBL2 regulates de novo purine synthesis by transcriptionally activating IMPDH1 in hepatocellular carcinoma cells.

BACKGROUND: Metabolic reprogramming is a hallmark of cancer, alteration of nucleotide metabolism of hepatocellular carcinoma (HCC) is not well-understood. MYBL2 regulates cell cycle progression and hepatocarcinogenesis, its role in metabolic regulation remains elusive. PATIENTS AND METHODS: Copy number, mRNA and protein level of MYBL2 and IMPDH1 were analyzed in HCC, and correlated with patient survival. Chromatin Immunoprecipitation sequencing (Chip-seq) and Chromatin Immunoprecipitation quantitative polymerase chain reaction (ChIP-qPCR) were used to explore the relationship between MYBL2 and IMPDH1. Metabolomics were used to analyze how MYBL2 affected purine metabolism. The regulating effect of MYBL2 in HCC was further validated in vivo using xenograft models. RESULTS: The Results showed that copy-number alterations of MYBL2 occur in about 10% of human HCC. Expression of MYBL2, IMPDH1, or combination of both were significantly upregulated and associated with poor prognosis in HCC. Correlation, ChIP-seq and ChIP-qPCR analysis revealed that MYBL2 activates transcription of IMPDH1, while knock-out of MYBL2 retarded IMPDH1 expression and inhibited proliferation of HCC cells. Metabolomic analysis post knocking-out of MYBL2 demonstrated that it was essential in de novo purine synthesis, especially guanine nucleotides. In vivo analysis using xenograft tumors also revealed MYBL2 regulated purine synthesis by regulating IMPDH1, and thus, influencing tumor progression. CONCLUSION: MYBL2 is a key regulator of purine synthesis and promotes HCC progression by transcriptionally activating IMPDH1, it could be a potential candidate for targeted therapy for HCC.