Cancer cells preferentially metabolize glucose by aerobic glycolysis, characterized by increased lactate production. This distinctive metabolism involves expression of the embryonic M2 isozyme of pyruvate kinase, in contrast to the M1 isozyme normally expressed in differentiated cells, and it confers a proliferative advantage to tumor cells. The M1 and M2 pyruvate-kinase isozymes are expressed from a single gene through alternative splicing of a pair of mutually exclusive exons. We measured the expression of M1 and M2 mRNA and protein isoforms in mouse tissues, tumor cell lines, and during terminal differentiation of muscle cells, and show that alternative splicing regulation is sufficient to account for the levels of expressed protein isoforms. We further show that the M1-specific exon is actively repressed in cancer-cell lines-although some M1 mRNA is expressed in cell lines derived from brain tumors -and demonstrate that the related splicing repressors hnRNP A1 and A2, as well as the polypyrimidine-tract-binding protein PTB, contribute to this control. Downregulation of these splicing repressors in cancer-cell lines using shRNAs rescues M1 isoform expression and decreases the extent of lactate production. These findings extend the links between alternative splicing and cancer, and begin to define some of the factors responsible for the switch to aerobic glycolysis.aerobic glycolysis | cancer | RNA splicing C ancer cells exhibit a metabolic phenotype characterized by increased glycolysis with lactate generation, regardless of oxygen availability-a phenomenon termed the Warburg effect. Recent work demonstrated that expression of the type II isoform of the pyruvate-kinase-M gene (PKM2, referred to here as PK-M) is a critical determinant of this metabolic phenotype, and confers a selective proliferative advantage to tumor cells in vivo (1). This finding adds to the growing body of evidence that alterations in alternative prem-RNA splicing play important roles in different aspects of cancer progression (2, 3).Pyruvate-kinase (PK) is the enzyme that catalyzes the final step in glycolysis, generating pyruvate and ATP from phosphoenolpyruvate and ADP (4). The resulting pyruvate can be converted to lactate or it can be incorporated into the tricarboxylic acid (TCA) cycle to drive oxidative phosphorylation. PK is encoded by two paralogous genes, each of which is alternatively spliced, such that four PK isoforms are expressed in mammals. The L and R isozymes, derived from the PKLR gene, show tissue-specific expression in the liver and red-blood cells, respectively. They have different first exons, defined by tissue-specific promoters (5). The PKM2 (PK-M) gene consists of 12 exons, of which exons 9 and 10 are alternatively spliced in a mutually exclusive fashion to give rise to the PK-M1 and PK-M2 isoforms, respectively (6). Exons 9 and 10 each encode a 56 amino acid variable segment that confers distinctive properties to the regulation and activity of PK-M1 and PK-M2 enzymes; as a result, PK-M1 is constitutively active,...

Alternative splicing of the pyruvate kinase M gene (PK-M) can generate the M2 isoform and promote aerobic glycolysis and tumor growth. However, the cancer-specific alternative splicing regulation of PK-M is not completely understood. Here, we demonstrate that PK-M is regulated by reciprocal effects on the mutually exclusive exons 9 and 10, such that exon 9 is repressed and exon 10 is activated in cancer cells. Strikingly, exonic, rather than intronic, cis-elements are key determinants of PK-M splicing isoform ratios. Using a systematic sub-exonic duplication approach, we identify a potent exonic splicing enhancer in exon 10, which differs from its homologous counterpart in exon 9 by only two nucleotides. We identify SRSF3 as one of the cognate factors, and show that this serine/arginine-rich protein activates exon 10 and mediates changes in glucose metabolism. These findings provide mechanistic insights into the complex regulation of alternative splicing of a key regulator of the Warburg effect, and also have implications for other genes with a similar pattern of alternative splicing.

Aims: 1 Describe the epidemiology and determine risk factors for COVID-19 associated mucormycosis. 2 Elaborate the clinical spectrum of Rhino-Orbital-Cerebral Mucormycosis (ROCM), pattern of neuroaxis involvement and it's radiological correlates. Methods Observational study. Consecutive, confirmed cases of mucormycosis (N = 55) were included. A case of mucormycosis was defined as one who had clinical and radiological features consistent with mucormycosis along with demonstration of the fungus in tissue via KOH mount/culture/histopathological examination (HPE). Data pertaining to epidemiology, risk factors, clinico-radiological features were analysed using percentage of total cases. Results Middle aged, diabetic males with recent COVID-19 infection were most affected. New onset upper jaw toothache was a striking observation in several cases. Among neurological manifestations headache, proptosis, vision loss, extraocular movement restriction; cavernous sinus, meningeal and parenchymal involvement were common. Stroke in ROCM followed a definitive pattern with watershed infarction. Conclusions New onset upper jaw toothache and loosening of teeth should prompt an immediate search for mucormycosis in backdrop of diabetic patients with recent COVID-19 disease, aiding earlier diagnosis and treatment initiation. Neuroaxis involvement was characterized by a multitude of features pertaining to involvement of optic nerve, extraocular muscles, meninges, brain parenchyma and internal carotid artery.

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