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Abstract Background Glycogen storage disease (GSD) type IXb is one of the rare variants of GSDs. It is a genetically heterogeneous metabolic disorder due to deficient hepatic phosphorylase kinase activity. Diagnosis of GSD can be difficult because of overlapping manifestations. Mutation analysis of the genes related to each type of GSD is supposed to be problem-solving, however, the presence of novel mutations can be confusing. In this case report, we will describe our experience with a young girl with the diagnosis of GSD and two novel mutations related to GSD type IXb. Case presentation A 3-year- old girl presented with short stature, hepatomegaly, and liver cirrhosis. No specific diagnosis was made based on laboratory data, so liver biopsy and targeted-gene sequencing (TGS) were performed to find out the specific molecular basis of her disease. It was confirmed that the patient carries two novel variants in the PHKB gene. The variant in the PHKB gene was classified as pathogenic. Conclusions This is the first reported case of a dual molecular mutation of glycogen storage disease type IXb in the same patient. Two novel variants in PHKB were identified and one of them was a pathogenic split-site mutation. In conclusion, for the first time, identification of the novel variants in this patient expands the molecular and the phenotype basis of dual variants in GSD-IXb.

Background: Glycogen storage disease (GSD) type IXb is one of the rare variants of GSDs. It is a genetically heterogeneous metabolic disorder due to deficient hepatic phosphorylase kinase activity. Diagnosis of GSD can be difficult because of overlapping manifestations. Mutation analysis of the genes related to each type of GSD is supposed to be problem-solving, however, the presence of novel mutations can be confusing. In this case report, we will describe our experience with a young girl with the diagnosis of GSD and two novel mutations related to GSD type IXb.Case Presentation: A 3-year- old girl presented with short stature, hepatomegaly, and liver cirrhosis. No specific diagnosis was made based on laboratory data, so liver biopsy and targeted-gene sequencing (TGS) were performed to find out the specific molecular basis of her disease. It was confirmed that the patient carries two novel variants in the PHKB gene. The variant in the PHKB gene was classified as pathogenic.Conclusions: This is the first reported case of a dual molecular mutation of glycogen storage disease type IXb in the same patient. Two novel variants in PHKB were identified and one of them was a pathogenic split-site mutation. In conclusion, for the first time, identification of the novel variants in this patient expands the molecular and the phenotype basis of dual variants in GSD-IXb. [ABSTRACT FROM AUTHOR]

Many functions of phosphorylase kinase (PhK) are regulated by Ca2+ and Mg2+ ions. Ca2+ and Mg2+ ions stimulate activity of PhK, induce the changes in the tertiary and quaternary structure of the hexadecameric enzyme molecule, provoke association/aggregation of PhK molecules, enhance PhK binding to glycogen. To establish the kinetic regime of Ca2+ and Mg2+-induced aggregation of PhK from rabbit skeletal muscles at 40 °C, in the present work the kinetics of aggregation was studied at various protein concentrations using the dynamic light scattering. The proposed mechanism of aggregation involves the stage of unfolding of the protein molecule with retention of the integrity of its oligomeric structure, the nucleation stage and stages of the growth of protein aggregates. The initial rate of the aggregation process at the stage of aggregate growth depends linearly on the protein concentration. This means that the order of aggregation with respect to the protein is equal to unity and the aggregation rate is limited by the rate of protein unfolding. The rate constant of the first order characterizing the stage of protein unfolding was found to be equal to 0.071 min−1 (40 mM Hepes, pH 6.8, 100 mM NaCl, 0.1 mM Ca2+, 10 mM Mg2+). Unlabelled Image • Many functions of phosphorylase kinase are regulated by Ca2+ and Mg2+ ions. • Kinetic regime of aggregation of highly complexly regulated protein is established. • The rate-limiting stage of the aggregation process is the stage of protein unfolding. • Aggregation pathway has been analyzed at various protein concentrations. • The mechanism of aggregation of phosphorylase kinase at 40 °C has been proposed. [ABSTRACT FROM AUTHOR]

Background: Necroptosis is a programmed necrotic cell death, in which dying cells rupture and release intracellular components that trigger a proinflammatory response. The current study aimed at probing the circular RNA (circRNA)-mediated regulatory mechanisms in necroptosis in premature ovarian failure (POF)., Methods: CircRNA sequencing analysis was conducted in ovarian tissues of control and POF rats and transcriptome microarrays were acquired from the GSE33423 dataset. Differential expression analysis of circRNAs and mRNAs was executed between the POF and control data. Both a necroptosis-based circRNA-microRNA (miRNA)-mRNA network and a protein-protein interaction (PPI) network were established. Then, the functional annotation and immunological traits were analyzed., Results: Totally, 1266 upregulated and 1283 downregulated circRNAs as well as 1101 upregulated and 1168 downregulated mRNAs were determined in the POF rats versus the controls. The differentially expressed mRNAs predominantly correlated with necroptosis. The circRNA-miRNA-mRNA networks of downregulated necroptosis genes (comprising rno_circRNA_004995-rno-miR-148b-5p-H2afy2, rno_circRNA_016998-rno-miR-29a-5p-Hmgb1, and rno_circRNA_017593-rno-miR-29a-5p-Hmgb1) and upregulated necroptosis genes (comprising rno_circRNA_015900-rno-miR-935-Stat1, rno_circRNA_007946-rno-miR-328a-3p-Stat5a, rno_circRNA_007947-rno-miR-328a-3p-Stat5a, rno_circRNA_005064-rno-miR-18a-5p-Stat1, rno_circRNA_005064-rno-miR-18a-5p-Stat5a, rno_circRNA_005115-rno-miR-22-3p-Stat1, rno_circRNA_009028-rno-miR-342-5p-Stat1, rno_circRNA_011240-rno-miR-1224-Stat5a, rno_circRNA_016078-rno-miR-711-Stat5a) were built. POF-specific necroptosis genes ( STAT1, STAT5A, PLA2G4A, HMG1L1, HMGB1, AGER, EGFR, HDAC7, IFNA1, IL10RB, IL27RA, PYGL, SOCS1, TRADD, CXCL10, DDX5, EZH2, FADS2, FER, H2AFY2, HIST1H2AF, IFI44L, IL27, IRGM, MX1, NFKB2, PAFAH2, PEMT, PGM2L1, PGR , PHKA2 , and PLB1 ) were selected since they displayed notable associations with most immune cells, immune checkpoints, chemokines, human leukocyte antigen (HLA) molecules, and immune receptors., Conclusions: Altogether, we proposed the presence of widespread regulatory mechanisms of circRNAs in necroptosis and demonstrated that altered circRNA biogenesis might contribute to POF by affecting necroptosis., Competing Interests: The authors declare no conflict of interest., (© 2023 The Author(s). Published by IMR Press.)

Background: PHKG2 gene mutation can lead to liver phosphorylase kinase (PhK) deficiency, which is related to glycogen storage disease type IX (GSD IX). GSD IXc due to PHKG2 mutation is the second most common GSD IX. Methods: We identified a novel mutation (c.553C>T, p.Arg185X) in PHKG2 in a Chinese family and verified it by next-generation and Sanger sequencing. The mutation spectrum of the PHKG2 gene was summarized based on 25 GSD IXc patients with PHKG2 mutations. Results: We found that missense mutation (39%) was the most common type of mutation, followed by nonsense mutation (23%). Mutations were more prevalent in Asian (12/25) and European (9/25) populations than in populations from elsewhere. The exons had more sites of mutation than the introns, and exons 3 and 6 were the most frequent sites of mutations. Conclusions: This study expands our knowledge of the PHKG2 gene mutation spectrum, providing a molecular basis for GSD IXc. [ABSTRACT FROM AUTHOR]

Abstract: Phosphorylase kinase (PhK), a 1.3 MDa regulatory enzyme complex in the glycogenolysis cascade, has four copies each of four subunits, (αβγδ)4, and 325 kDa of unique sequence (the mass of an αβγδ protomer). The α, β and δ subunits are regulatory, and contain allosteric activation sites that stimulate the activity of the catalytic γ subunit in response to diverse signaling molecules. Due to its size and complexity, no high resolution structures have been solved for the intact complex or its regulatory α and β subunits. Of PhK's four subunits, the least is known about the structure and function of its largest subunit, α. Here, we have modeled the full‐length α subunit, compared that structure against previously predicted domains within this subunit, and performed hydrogen‐deuterium exchange on the intact subunit within the PhK complex. Our modeling results show α to comprise two major domains: an N‐terminal glycoside hydrolase domain and a large C‐terminal importin α/β‐like domain. This structure is similar to our previously published model for the homologous β subunit, although clear structural differences are present. The overall highly helical structure with several intervening hinge regions is consistent with our hydrogen‐deuterium exchange results obtained for this subunit as part of the (αβγδ)4 PhK complex. Several low exchanging regions predicted to lack ordered secondary structure are consistent with inter‐subunit contact sites for α in the quaternary structure of PhK; of particular interest is a low‐exchanging region in the C‐terminus of α that is known to bind the regulatory domain of the catalytic γ subunit. [ABSTRACT FROM AUTHOR]

Abstract: In the tightly regulated glycogenolysis cascade, the breakdown of glycogen to glucose‐1‐phosphate, phosphorylase kinase (PhK) plays a key role in regulating the activity of glycogen phosphorylase. PhK is a 1.3 MDa hexadecamer, with four copies each of four different subunits (α, β, γ and δ), making the study of its structure challenging. Using hydrogen‐deuterium exchange, we have analyzed the regulatory β subunit and the catalytic γ subunit in the context of the intact non‐activated PhK complex to study the structure of these subunits and identify regions of surface exposure. Our data suggest that within the non‐activated complex the γ subunit assumes an activated conformation and are consistent with a previous docking model of the β subunit within the cryoelectron microscopy envelope of PhK. [ABSTRACT FROM AUTHOR]

Glycogen storage disease (GSD) type IXa is caused by PHKA2 mutation, which accounts for about 75% of all the GSD type IX cases. Here we first summarized the clinical data and analyzed the PHKA2 gene of 17 Chinese male patients suspected of having GSD type IXa. Clinical symptoms of our patients included hepatomegaly, growth retardation, and liver dysfunction. The clinical and biochemical manifestations improved and even disappeared with age. We detected 14 mutations in 17 patients, including 8 novel mutations; exons 2 and 4 were hot spots in this research. In conclusion, glycogen storage disease type IXa is a mild disorder with a favorable prognosis, and there was no relationship between genotype and phenotype of this disease. [ABSTRACT FROM AUTHOR]

Background: When ectopically overexpressed, anticancer genes, such as TRAIL, PAR4 and ORCTL3, specifically destroy tumour cells without harming untransformed cells. Anticancer genes can not only serve as powerful tumour specific therapy tools but studying their mode of action can reveal mechanisms underlying the neoplastic transformation, sustenance and spread., Methods: Anticancer gene discovery is normally accidental. Here we describe a systematic, gain of function, forward genetic screen in mammalian cells to isolate novel anticancer genes of human origin. Continuing with over 30,000 transcripts from our previous study, 377 cell death inducing genes were subjected to screening. FBLN5 was chosen, as a proof of principle, for mechanistic gene expression profiling, comparison pathways analyses and functional studies., Results: Sixteen novel anticancer genes were isolated; these included non-coding RNAs, protein-coding genes and novel transcripts, such as ZNF436-AS1, SMLR1, TMEFF2, LINC01529, HYAL2, NEIL2, FBLN5, YPEL4 and PHKA2-processed transcript. FBLN5 selectively caused inhibition of MYC in COS-7 (transformed) cells but not in CV-1 (normal) cells. MYC was identified as synthetic lethality partner of FBLN5 where MYC transformed CV-1 cells experienced cell death upon FBLN5 transfection, whereas FBLN5 lost cell death induction in MCF-7 cells upon MYC knockdown., Conclusions: Sixteen novel anticancer genes are present in human genome including FBLN5. MYC is a synthetic lethality partner of FBLN5. Video Abstract., (© 2023. BioMed Central Ltd., part of Springer Nature.)

X-linked liver glycogenosis (XLG), also known as glycogen storage disease type-IXa, is characterized by hepatomegaly, abnormal liver functions and growth retardation. It is caused by mutations in the PHKA2 gene that encodes the α-subunit of phosphorylase kinase (PHK). XLG can be divided into two subtypes: XLG-I, with a deficiency in PHK activity in peripheral blood cells and the liver; and XLG-II, with normal PHK activity in vitro. This report describes two boys who presented with hepatomegaly and abnormal liver function. Pedigree analysis revealed them to be fifth-degree relatives, with the disease transmitted through undiagnosed grandfathers. Liver histology confirmed GSD diagnosis, and both cases had a deficiency in PHK activity in red blood cells and liver tissues. This is the first report of XLG-I in the ethnic-Chinese population in Taiwan. This report indicates that XLG may be undiagnosed or underestimated. A correct diagnosis is necessary for proper management and genetic counseling.

Ferroptosis is a newly discovered form of regulatory cell death induced by iron-dependent lipid peroxidation. Infection with Helicobacter pylori (H. pylori) is regarded as a high-risk factor for the development of gastric cancer (GC) and is associated with an increase in the levels of reactive oxygen species with activation of oncogenic signaling pathways. However, whether GC arising in the context of infection with H. pylori is correlated with ferroptosis is still unknown. In this study, we demonstrate that H. pylori infection increased the sensitivity of GC cells to RSL3 (RAS-selective lethal3)-induced ferroptosis. The molecular subtypes mediated by ferroptosis-related genes are associated with tumor microenvironment (TME) cell infiltration and patient survival. Importantly, we identified that the expression of phosphorylase kinase G2 (PHKG2) was remarkably correlated with H. pylori infection, metabolic biological processes, patient survival and therapy response. We further found the mechanism of H. pylori-induced cell sensitivity to ferroptosis, which involves PHKG2 regulation of the lipoxygenase enzyme Arachidonate 5-Lipoxygenase (ALOX5). In conclusion, PHKG2 facilitates RSL3-induced ferroptosis in H. pylori-positive GC cells by promoting ALOX5 expression. These findings may contribute to a better understanding of the unique pathogenesis of H. pylori-induced GC and allow for maximum efficacy of genetic, cellular, and immune therapies for controlling ferroptosis in diverse contexts., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)