A deficiency of striated preferentially expressed gene (Speg), a member of the myosin light chain kinase family, results in abnormal myofibril structure and function of immature cardiomyocytes (CMs), corresponding with a dilated cardiomyopathy, heart failure and perinatal death. Mitochondrial development plays a role in cardiomyocyte maturation. Therefore, this study investigated whether Speg deficiency ( - / - ) in CMs would result in mitochondrial abnormalities. Speg wild-type and Speg-/- C57BL/6 littermate mice were utilized for assessment of mitochondrial structure by transmission electron and confocal microscopies. Speg was expressed in the first and second heart fields at embryonic (E) day 7.5, prior to the expression of mitochondrial Na+/Ca2+/Li+ exchanger (NCLX) at E8.5. Decreases in NCLX expression (E11.5) and the mitochondrial-to-nuclear DNA ratio (E13.5) were observed in Speg-/- hearts. Imaging of E18.5 Speg-/- hearts revealed abnormal mitochondrial cristae, corresponding with decreased ATP production in cells fed glucose or palmitate, increased levels of mitochondrial superoxide and depolarization of mitochondrial membrane potential. Interestingly, phosphorylated (p) PGC-1α, a key mediator of mitochondrial development, was significantly reduced in Speg-/- hearts during screening for targeted genes. Besides Z-line expression, Speg partially co-localized with PGC-1α in the sarcomeric region and was found in the same complex by co-immunoprecipitation. Overexpression of a Speg internal serine/threonine kinase domain in Speg-/- CMs promoted translocation of pPGC-1α into the nucleus, and restored ATP production that was abolished by siRNA-mediated silencing of PGC-1α. Our results demonstrate a critical role of Speg in mitochondrial development and energy metabolism in CMs, mediated in part by phosphorylation of PGC-1α.

UNASSIGNED: Through transcriptomic and metabolomic analyses, this study examined the role of high-fiber diet in obesity complicated by diabetes and neurodegenerative symptoms.
UNASSIGNED: The expression matrix of high-fiber-diet-related metabolites, blood methylation profile associated with pre-symptomatic dementia in elderly patients with type 2 diabetes mellitus (T2DM), and high-throughput single-cell sequencing data of hippocampal samples from patients with Alzheimer\'s disease (AD) were retrieved from the Gene Expression Omnibus (GEO) database and through a literature search. Data were analyzed using principal component analysis (PCA) after quality control and data filtering to identify different cell clusters and candidate markers. A protein-protein interaction network was mapped using the STRING database. To further investigate the interaction among high-fiber-diet-related metabolites, methylation-related DEGs related to T2DM, and single-cell marker genes related to AD, AutoDock was used for semi-flexible molecular docking.
UNASSIGNED: Based on GEO database data and previous studies, 24 marker genes associated with high-fiber diet, T2DM, and AD were identified. Top 10 core genes include SYNE1, ANK2, SPEG, PDZD2, KALRN, PTPRM, PTPRK, BIN1, DOCK9, and NPNT, and their functions are primarily related to autophagy. According to molecular docking analysis, acetamidobenzoic acid, the most substantially altered metabolic marker associated with a high-fiber diet, had the strongest binding affinity for SPEG.
UNASSIGNED: By targeting the SPEG protein in the hippocampus, acetamidobenzoic acid, a metabolite associated with high-fiber diet, may improve diabetic and neurodegenerative diseases in obese people.

The coronavirus disease 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has led to more than 6.4 million deaths worldwide. The prevalent comorbidity between hypertension and severe COVID-19 suggests common genetic factors may affect the outcome of both diseases. As both hypertension and severe COVID-19 demonstrate sex-biased prevalence, common genetic factors between the two diseases may display sex-biased differential associations. By evaluating COVID-19 association signals of 172-candidate hypertension single nucleotide polymorphisms (SNPs) derived from more than 1 million European individuals in two sex-stratified severe COVID-19 genome-wide association studies from UK BioBank with European ancestry, we revealed one functional cis expression quantitative trait locus of SPEG (rs12474050) showing sex-biased association with severe COVID-19 in women. The risk allele rs12474050*T associates with higher blood pressure. In our study, we found it is significantly correlated with lower SPEG expression in muscle-skeletal but with higher expression in both brain cerebellum and cerebellar hemisphere. Additionally, nominal significances were detected for the association between rs12474050*T and lower SPEG expression in both heart left ventricle and atrial appendage; among these tissues, the SPEG expression is nominally significantly higher in females than in males. Further analysis revealed SPEG is mainly expressed in cardiomyocytes in heart and is upregulated upon SARS-CoV-2 infection, with significantly higher upregulation of SPEG only observed in female but not in male COVID-19 patients compared to both normal female and male individuals, suggesting upregulation of SPEG is a female-specific protective mechanism against COVID-19 induced heart damage. Taken together, our analyses suggest the involvement of SPEG in both hypertension and severe COVID-19 in women, which provides new insights for sex-biased effect of severe COVID-19 in women.

Spermidine and other polyamines alleviate oxidative stress, yet excess spermidine seems toxic to Escherichia coli unless it is neutralized by SpeG, an enzyme for the spermidine N-acetyl transferase function. Thus, wild-type E. coli can tolerate applied exogenous spermidine stress, but ΔspeG strain of E. coli fails to do that. Here, using different reactive oxygen species (ROS) probes and performing electron paramagnetic resonance spectroscopy, we provide evidence that although spermidine mitigates oxidative stress by lowering overall ROS levels, excess of it simultaneously triggers the production of superoxide radicals, thereby causing toxicity in the ΔspeG strain. Furthermore, performing microarray experiment and other biochemical assays, we show that the spermidine-induced superoxide anions affected redox balance and iron homeostasis. Finally, we demonstrate that while RNA-bound spermidine inhibits iron oxidation, free spermidine interacts and oxidizes the iron to evoke superoxide radicals directly. Therefore, we propose that the spermidine-induced superoxide generation is one of the major causes of spermidine toxicity in E. coli.

Centronuclear myopathy (CNM) is a congenital neuromuscular disorder caused by pathogenic variation in genes associated with membrane trafficking and excitation-contraction coupling (ECC). Bi-allelic autosomal-recessive mutations in striated muscle enriched protein kinase (SPEG) account for a subset of CNM patients. Previous research has been limited by the perinatal lethality of constitutive Speg knockout mice. Thus, the precise biological role of SPEG in developing skeletal muscle remains unknown. To address this issue, we generated zebrafish spega, spegb and spega;spegb (speg-DKO) mutant lines. We demonstrated that speg-DKO zebrafish faithfully recapitulate multiple phenotypes associated with CNM, including disruption of the ECC machinery, dysregulation of calcium homeostasis during ECC and impairment of muscle performance. Taking advantage of zebrafish models of multiple CNM genetic subtypes, we compared novel and known disease markers in speg-DKO with mtm1-KO and DNM2-S619L transgenic zebrafish. We observed Desmin accumulation common to all CNM subtypes, and Dnm2 upregulation in muscle of both speg-DKO and mtm1-KO zebrafish. In all, we establish a new model of SPEG-related CNM, and identify abnormalities in this model suitable for defining disease pathomechanisms and evaluating potential therapies. This article has an associated First Person interview with the joint first authors of the paper.

Congenital myopathies are a heterogeneous group of conditions diagnosed based on the clinical presentation, muscle histopathology and genetic defects. Recessive mutations in the SPEG gene have been described in recent years and are primarily associated with centronuclear myopathy with cardiomyopathy. In this report, we describe two Brazilian siblings, aged 13 and 6 years, with a novel homozygous mutation (c.8872 C>T:p.Arg2958Ter) in the SPEG gene leading to a congenital myopathy. In the older sibling, the muscle biopsy showed fiber size disproportion. The mean diameter of type 2 fibers (119 µm) was significantly higher than type 1 (57 µm) (P < 0,001) with a 72% prevalence of type 1 fibers. The patient also had progressive cardiomyopathy treated with heart transplantation. The present report expands the muscle histopathological findings related to mutations in the SPEG gene, including fiber size disproportion without central nuclei. Additionally, this report describes the first case of heart transplantation in a patient with SPEG mutations.

Centronuclear myopathies (CNMs) are a subgroup of congenital myopathies (CMs) characterized by muscle weakness, genetic heterogeneity, and predominant type 1 fibers and increased central nuclei in muscle biopsy. Mutations in CNM-causing genes such as MTM1, DNM2, BIN1, RYR1, CACNA1S, TTN, and extraordinary rarely SPEG (striated muscle preferentially expressed protein kinase) have been identified for about 60-80% of patients. Herein, we report a case of CM due to a novel variation in the SPEG gene, manifested by mild neonatal hypotonia, muscle weakness, delayed motor milestones, and ophthalmoplegia, without dilated cardiomyopathy. We identified a novel variation [c.153C>T (p.Asn51=) in exon 1] in the SPEG gene with whole-exome sequencing and confirmed by Sanger sequencing. Mild intellectual disability has not been associated with SPEG-related CM in the previous reports. We suggest that this report expands the phenotypic spectrum of SPEG-related CM, and further case reports are required to expand the genotype-phenotype correlations.

Aberrant glycosylation has long been known to be associated with cancer, since it is involved in key mechanisms such as tumour onset, development and progression. This review will focus on protein glycosylation studies in cells, tissue, urine and serum in the context of prostate cancer. A dedicated section will cover the glycoforms of prostate specific antigen, the molecule that, despite some important limitations, is routinely tested for helping prostate cancer diagnosis. Our aim is to provide readers with an overview of mass spectrometry-based glycoproteomics of prostate cancer. From this perspective, the first part of this review will illustrate the main strategies for glycopeptide enrichment and mass spectrometric analysis. The molecular information obtained by glycoproteomic analysis performed by mass spectrometry has led to new insights into the mechanism linking aberrant glycosylation to cancer cell proliferation, migration and immunoescape.

The SpeG spermidine/spermine N-acetyltransferase (SSAT) from Escherichia coli belongs to the Gcn5-related N-acetyltransferase (GNAT) superfamily of proteins. In vitro characterization of this enzyme shows it acetylates the polyamines spermine and spermidine, with a preference toward spermine. This enzyme has a conserved tyrosine residue (Y135) that is found in all SSAT proteins and many GNAT functional subfamilies. It is located near acetyl coenzyme A in the active center of these proteins and has been suggested to act as a general acid in a general acid/base chemical mechanism. In contrast, a previous study showed this residue was not critical for E. coli SpeG enzymatic activity when mutated to phenylalanine. This result was quite different from previous studies with a comparable residue in the human and mouse SSAT proteins, which also acetylate spermine and spermidine. Therefore, we constructed several mutants of the E. coli SpeG Y135 residue and tested their enzymatic activity. We found this conserved residue was indeed critical for E. coli SpeG enzyme activity and may behave similarly in other SSAT proteins.

Three members of the obscurin protein family that contain tandem kinase domains with important signaling functions for cardiac and striated muscles are the giant protein obscurin, its obscurin-associated kinase splice isoform, and the striated muscle enriched protein kinase (SPEG). While there is increasing evidence for the specific roles that each individual kinase domain plays in cross-striated muscles, their biology and regulation remains enigmatic. Our present study focuses on kinase domain 1 and the adjacent low sequence complexity inter-kinase domain linker in obscurin and SPEG. Using Phos-tag gels, we show that the linker in obscurin contains several phosphorylation sites, while the same region in SPEG remained unphosphorylated. Our homology modeling, mutational analysis and molecular docking demonstrate that kinase 1 in obscurin harbors all key amino acids important for its catalytic function and that actions of this domain result in autophosphorylation of the protein. Our bioinformatics analyses also assign a list of putative substrates for kinase domain 1 in obscurin and SPEG, based on the known and our newly proposed phosphorylation sites in muscle proteins, including obscurin itself.