Squalene synthase (farnesyl-diphosphate:farnesyl-diphosphate farnesyltransferase, EC 2.5.1.21) is the first enzyme specific to the cholesterol biosynthetic pathway. The activity of rat hepatic squalene synthase is regulated by dietary cholesterol and by the dietary 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) 1 reductase inhibitors, lovastatin, or fluvastatin (1, 2). The activity of human squalene synthase (HSS) and the level of its mRNA are regulated by sterols in the human hepatoma cell line HepG2. Sterol-mediated regulation has been localized to a 10-base pair (bp) element in the 5Ј-flanking region of other sterolregulated genes. This 10-bp sterol regulatory element 1 (SRE-1) mediates increased transcription of the genes encoding HMG-CoA synthase and the low density lipoprotein (LDL) receptor in sterol-depleted cells, and its activity is inhibited by sterols (3, 4). Proteins that bind to the SRE-1 of the LDL receptor (SREBPs) were purified by DNA affinity chromatography from nuclear extracts of HeLa cells. A cDNA for SREBP-1 was isolated from adipocyte cDNA library (5). This cDNA, designated ADD1, activated transcription of a reporter gene containing an "E-box" sequence present in the promoter of fatty acid synthase in transfected NIH 3T3 cells. Cloned SREBP cDNA contain two major classes of proteins, SREBP-1 (5) and SREBP-2 (8). Three different cDNAs for SREBP-1 were isolated, suggesting multiple forms of the mRNA and perhaps different proteins as well. The physiological significance of these subclasses is unclear (6). Different SREBP-1 proteins may have specific physiological roles because mRNAs for the various isoforms are differentially regulated by sterol depletion in HepG2 cells (8).Proteolytic cleavage of the C-terminal membrane-associated domain of the nascent SREBP-1 (125 kDa) forms its nuclear form (68 kDa). This proteolytic maturation was proposed to be accomplished by a sterol-inhibited protease. The calpain inhibitor N-acetyl-leucyl-leucyl-norleucinal (ALLN) induced the mRNA for HMG-CoA synthase and was proposed to inhibit the degradation of the mature SREBP-1 (9).In other sterol-regulated genes, the SRE-1 is not involved in sterol-mediated transcriptional regulation. Although the promoter region of farnesyl diphosphate synthase contains multiple forms of the SRE-1 element, these elements are not involved in the sterol-mediated transcriptional regulation (10). Similarly, the promoter of the hamster HMG-CoA reductase contains unique sites for sterol regulation. Red 25, a nuclear hamster liver protein, binds to this regulatory region but did not bind to the sterol regulatory regions of the LDL receptor and HMG-CoA synthase promoters (11).In this report we characterize the 5Ј region of the HSS gene. The promoter activity and the sterol-mediated regulation of this DNA were assessed by fusing 5Ј-flanking DNA to a luciferase reporter gene and transfecting it into HepG2 cells and Chinese hamster ovary (CHO-K1) cells. A 69 bp DNA sequence confers transcriptional competence and sterol regulation. ADD1...

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Cholinesterases are serine esterases that rapidly hydrolyze the neurotransmitter acetylcholine. In humans, cholinesterases exhibit extensive polymorphism in terms of their substrate specificity, sensitivity to selective inhibitors, hydrophobicity, and cellular as well as subcellular localization. It is not yet known whether the various cholinesterase forms originate from different genes or are products of posttranscriptional and posttranslational processing. The extent to which these enzyme forms are homologous in their amino acid sequence is also not known. However, a consensus organophosphate-binding hexapeptide sequence Phe-Gly-Glu-Ser-Ala-Gly was found both in "true" acetylcholinesterase from the electric organ of Torpedo [McPhee-Quigley et al: J Biol Chem 260:12185-12189, 1985] and in "pseudocholinesterase" (butyrylcholinesterase) from human serum [Lockridge: "Cholinesterases--Fundamental and Applied Aspects." New York: de Gruyter pp 5-12, 1984], suggesting that this region in the protein is conserved in all cholinesterases. Based on this common sequence, we prepared synthetic oligodeoxynucleotides and used them as labeled probes to screen a cDNA library from fetal human brain mRNA, cloned in lambda gt10 phages. A cDNA clone of 770 nucleotides in length was isolated. It contains an open reading frame terminating with the sequence Ser-Val-Thr-Leu-Phe-Gly-Glu-Ser-Ala-Gly-Ala-Ala, which includes the consensus hexapeptide used for designing the DNA probe. Furthermore, the sequence of this 12-amino acid peptide is identical to the sequence reported for the organophosphate binding site of human serum pseudocholinesterase [Lockridge: "Cholinesterases--Fundamental and Applied Aspects." New York: de Gruyter, pp 5-12, 1984]. These findings confirm that the isolated clone is indeed part of a human cholinesterase cDNA.

Purpose: Glycogen storage disease type IV (GSD IV) has historically been divided into discrete hepatic (classic hepatic, non-progressive hepatic) and neuromuscular (perinatal-congenital neuromuscular, juvenile neuromuscular) subtypes. However, the extent to which this subtype-based classification system accurately captures the landscape of phenotypic variation among GSD IV patients has not been systematically assessed.Methods: This study synthesized clinical data from all eligible cases of GSD IV in the published literature to evaluate whether this disorder is better conceptualized as discrete subtypes or a clinical continuum. A novel phenotypic scoring approach was applied to characterize the extent of hepatic, neuromuscular, and cardiac involvement in each eligible patient.Results: 146 patients met all inclusion criteria. The majority (61%) of those with sufficient data to be scored exhibited phenotypes that were not fully consistent with any of the established subtypes. These included patients who exhibited combined hepatic-neuromuscular involvement; patients whose phenotypes were intermediate between the established hepatic or neuromuscular subtypes; and patients who presented with predominantly cardiac disease.Conclusion: The application of this novel phenotypic scoring approach showed that–in contrast to the traditional subtype-based view–GSD IV may be better conceptualized as a multidimensional clinical continuum, whereby hepatic, neuromuscular, and cardiac involvement occur to varying degrees in different patients.

Glycogen storage disorders (GSDs) are inherited disorders of metabolism resulting from the deficiency of individual enzymes involved in the synthesis, transport, and degradation of glycogen. This literature review summarizes the development of gene therapy for the GSDs. The abnormal accumulation of glycogen and deficiency of glucose production in GSDs lead to unique symptoms based upon the enzyme step and tissues involved, such as liver and kidney involvement associated with severe hypoglycemia during fasting and the risk of long‐term complications including hepatic adenoma/carcinoma and end stage kidney disease in GSD Ia from glucose‐6‐phosphatase deficiency, and cardiac/skeletal/smooth muscle involvement associated with myopathy +/− cardiomyopathy and the risk for cardiorespiratory failure in Pompe disease. These symptoms are present to a variable degree in animal models for the GSDs, which have been utilized to evaluate new therapies including gene therapy and genome editing. Gene therapy for Pompe disease and GSD Ia has progressed to Phase I and Phase III clinical trials, respectively, and are evaluating the safety and bioactivity of adeno‐associated virus vectors. Clinical research to understand the natural history and progression of the GSDs provides invaluable outcome measures that serve as endpoints to evaluate benefits in clinical trials. While promising, gene therapy and genome editing face challenges with regard to clinical implementation, including immune responses and toxicities that have been revealed during clinical trials of gene therapy that are underway. Gene therapy for the glycogen storage diseases is under development, addressing an unmet need for specific, stable therapy for these conditions.