Identification of two novel ACAT1 variant associated with beta-ketothiolase deficiency in a 9-month-old boy.

OBJECTIVES: Mitochondrial acetoacetyl-CoA thiolase (beta-ketothiolase, T2) is necessary for the catabolism of ketone bodies andisoleucine. T2 deficiency is an autosomal recessive metabolic disorder caused by variant in the ACAT1 gene. In this report, we describe two novel ACAT1 variant identified in a Chinese family. CASE PRESENTATION: The 9-month-old male proband was admitted to the pediatric intensive care unit for altered consciousness. At the time of admission, the patient had acidosis, drowsiness, and respiratory failure. Both urine organic acid analyses and LC-MS/MS suggested T2 deficiency. Novel compound heterozygous variant (c.871G>C and c.1016_1017del) in the ACAT1 gene were detected in the proband by WES and verified through direct sequencing. Family analysis demonstrated that the first variant was transmitted from his father and the second variant was from his mother, indicating autosomal recessive inheritance. This report is the first to describe the association of these variant with T2 deficiency based on genetic testing. Although these variant were identified in the patient's elder sister and elder brother, they continue to be asymptomatic. CONCLUSIONS: We identified two novel ACAT1 variants associated with T2 deficiency. The identification expands the spectrum of known variant linked to the disorder.

Impact of various ß-ketothiolase genes on PHBHHx production in Cupriavidus necator H16 derivatives.

Poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] (PHBHHx) is a type of biopolyester of the polyhydroxyalkanoate group (PHA). Due to a wide range of properties resulting from the alteration of the (R)-3-hydroxyhexanoate (3HHx) composition, PHBHHx is getting a lot of attention as a substitute to conventional plastic materials for various applications. Cupriavidus necator H16 is the most promising PHA producer and has been genetically engineered to produce PHBHHx efficiently for many years. Nevertheless, the role of individual genes involved in PHBHHx biosynthesis is not well elaborated. C. necator H16 possesses six potential physiologically active ß-ketothiolase genes identified by transcriptome analysis, i.e., phaA, bktB, bktC (h16_A0170), h16_A0462, h16_A1528, and h16_B0759. In this study, we focused on the functionality of these genes in vivo in relation to 3HHx monomer supply. Gene deletion experiments identified BktB and H16_A1528 as important ß-ketothiolases for C6 metabolism in ß-oxidation. Furthermore, in the bktB/h16_A1528 double-deletion strain, the proportion of 3HHx composition of PHBHHx produced from sugar was very low, whereas that from plant oil was significantly higher. In fact, the proportion reached 36.2 mol% with overexpression of (R)-specifc enoyl-CoA hydratase (PhaJ) and PHA synthase. Furthermore, we demonstrated high-density production (196 g/L) of PHBHHx with high 3HHx (32.5 mol%) by fed-batch fermentation with palm kernel oil. The PHBHHx was amorphous according to the differential scanning calorimetry analysis. KEY POINTS: • Role of six ß-ketothiolases in PHBHHx biosynthesis was investigated in vivo. • Double-deletion of bktB/h16_A1528 results in high 3HHx composition with plant oil. • Amorphous PHBHHx with 32.5 mol% 3HHx was produced in high density by jar fermenter.

A novel missense variant in ACAA1 contributes to early-onset Alzheimer's disease, impairs lysosomal function, and facilitates amyloid-ß pathology and cognitive decline.

Alzheimer's disease (AD) is characterized by progressive synaptic dysfunction, neuronal death, and brain atrophy, with amyloid-ß (Aß) plaque deposits and hyperphosphorylated tau neurofibrillary tangle accumulation in the brain tissue, which all lead to loss of cognitive function. Pathogenic mutations in the well-known AD causal genes including APP, PSEN1, and PSEN2 impair a variety of pathways, including protein processing, axonal transport, and metabolic homeostasis. Here we identified a missense variant rs117916664 (c.896T>C, p.Asn299Ser [p.N299S]) of the acetyl-CoA acyltransferase 1 (ACAA1) gene in a Han Chinese AD family by whole-genome sequencing and validated its association with early-onset familial AD in an independent cohort. Further in vitro and in vivo evidence showed that ACAA1 p.N299S contributes to AD by disturbing its enzymatic activity, impairing lysosomal function, and aggravating the Aß pathology and neuronal loss, which finally caused cognitive impairment in a murine model. Our findings reveal a fundamental role of peroxisome-mediated lysosomal dysfunction in AD pathogenesis.

OCT1 - a yeast mitochondrial thiolase involved in the 3-oxoadipate pathway.

The 3-oxoacyl-CoA thiolases catalyze the last step of the fatty acid ß-oxidation pathway. In yeasts and plants, this pathway takes place exclusively in peroxisomes, whereas in animals it occurs in both peroxisomes and mitochondria. In contrast to baker's yeast Saccharomyces cerevisiae, yeast species from the Debaryomycetaceae family also encode a thiolase with predicted mitochondrial localization. These yeasts are able to utilize a range of hydroxyaromatic compounds via the 3-oxoadipate pathway the last step of which is catalyzed by 3-oxoadipyl-CoA thiolase and presumably occurs in mitochondria. In this work, we studied Oct1p, an ortholog of this enzyme from Candida parapsilosis. We found that the cells grown on a 3-oxoadipate pathway substrate exhibit increased levels of the OCT1 mRNA. Deletion of both OCT1 alleles impairs the growth of C. parapsilosis cells on 3-oxoadipate pathway substrates and this defect can be rescued by expression of the OCT1 gene from a plasmid vector. Subcellular localization experiments and LC-MS/MS analysis of enriched organellar fraction-proteins confirmed the presence of Oct1p in mitochondria. Phylogenetic profiling of Oct1p revealed an intricate evolutionary pattern indicating multiple horizontal gene transfers among different fungal groups.

Complete genome sequence of Photobacterium ganghwense C2.2: A new polyhydroxyalkanoate production candidate.

Polyhydroxyalkanoates (PHAs) are biodegradable bioplastics that can be manufactured sustainably and represent a promising green alternative to petrochemical-based plastics. Here, we describe the complete genome of a new marine PHA-producing bacterium-Photobacterium ganghwense (strain C2.2), which we have isolated from the Black Sea seashore. This new isolate is psychrotolerant and accumulates PHA when glycerol is provided as the main carbon source. Transmission electron microscopy, specific staining with Nile Red visualized via epifluorescence microscopy and gas chromatography analysis confirmed the accumulation of PHA. This is the only PHA-producing Photobacterium for which we now have a complete genome sequence, allowing us to investigate the pathways for PHA production and other secondary metabolite synthesis pathways. The de novo assembly genome, obtained using open-source tools, comprises two chromosomes (3.5, 2 Mbp) and a megaplasmid (202 kbp). We identify the entire PHA synthesis gene cluster that encodes a class I PHA synthase, a phasin, a 3-ketothiolase, and an acetoacetyl-CoA reductase. No conventional PHA depolymerase was identified in strain C2.2, but a putative lipase with extracellular amorphous PHA depolymerase activity was annotated, suggesting that C2.2 is unable to degrade intracellular PHA. A complete pathway for the conversion of glycerol to acetyl-CoA was annotated, in accordance with its ability to convert glycerol to PHA. Several secondary metabolite biosynthetic gene clusters and a low number of genes involved in antibiotic resistance and virulence were also identified, indicating the strain's suitability for biotechnological applications.

[Analysis of ACAT1 gene variants in a patient with ß-ketothiolase deficiency].

OBJECTIVE: To explore the genetic etiology of a child suspected for ß-ketothiolase deficiency by neonatal screening. METHODS: All coding exons and flanking sequences of the ACAT1 gene were subjected to targeted capture and high-throughput sequencing. Suspected variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The child was found to harbor compound heterozygous variants of the ACAT1 gene, namely c.121-3C>G and c.275G>A (p. Gly92Asp). The c.121-3C>G variant was also detected in his father and two sisters, while the c.275G>A (p. Gly92Asp) was a de novo variant. A c.334+ 172C>G (rs12226047) polymorphism was also detected in his mother and two sisters. Sanger sequencing has verified that the c.275G>A (p. Gly92Asp) and c.334+172C>G (rs12226047) variants are located on the same chromosome. Bioinformatics analysis suggested both c.121-3C>G and c.275G>A (p.G92D) variants to be damaging. Based on the American College of Medical Genetics and Genomics standards and guidelines, the c.275G>A variant of the ACAT1 gene was predicted to be pathogenic (PS2+ PM2+ PM3+ PP3+PP4), the c.121-3C>G variant to be likely pathogenic (PM2+ PM3+ PP3+PP4). CONCLUSION: The c.121-3C>G and c.275G>A variants of the ACAT1 gene probably underlay the pathogenesis of the child. Above finding has enriched the variant spectrum of the ACAT1 gene.

Analysis of ACAT1 gene variants in a patient with β-ketothiolase deficiency / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic etiology of a child suspected for β-ketothiolase deficiency by neonatal screening.@*METHODS@#All coding exons and flanking sequences of the ACAT1 gene were subjected to targeted capture and high-throughput sequencing. Suspected variants were verified by Sanger sequencing and bioinformatic analysis.@*RESULTS@#The child was found to harbor compound heterozygous variants of the ACAT1 gene, namely c.121-3C>G and c.275G>A (p. Gly92Asp). The c.121-3C>G variant was also detected in his father and two sisters, while the c.275G>A (p. Gly92Asp) was a de novo variant. A c.334+ 172C>G (rs12226047) polymorphism was also detected in his mother and two sisters. Sanger sequencing has verified that the c.275G>A (p. Gly92Asp) and c.334+172C>G (rs12226047) variants are located on the same chromosome. Bioinformatics analysis suggested both c.121-3C>G and c.275G>A (p.G92D) variants to be damaging. Based on the American College of Medical Genetics and Genomics standards and guidelines, the c.275G>A variant of the ACAT1 gene was predicted to be pathogenic (PS2+ PM2+ PM3+ PP3+PP4), the c.121-3C>G variant to be likely pathogenic (PM2+ PM3+ PP3+PP4).@*CONCLUSION@#The c.121-3C>G and c.275G>A variants of the ACAT1 gene probably underlay the pathogenesis of the child. Above finding has enriched the variant spectrum of the ACAT1 gene.

Cloning of wheat keto-acyl thiolase 2B reveals a role of jasmonic acid in grain weight determination.

Grain weight (GW) is one of the component traits of wheat yield. Existing reports have shown that multiple phytohormones are involved in the regulation of GW in different crops. However, the potential role of jasmonic acid (JA) remains unclear. Here, we report that triticale grain weight 1 (tgw1) mutant, with marked reductions in both GW and JA content, is caused by a premature stop mutation in keto-acyl thiolase 2B (KAT-2B) involved in ß-oxidation during JA synthesis. KAT-2B overexpression increases GW in wild type and boosts yield. Additionally, KAT-2B compliments the grain defect in tgw1 and rescues the lethal phenotype of the Arabidopsis kat2 mutant in a sucrose-free medium. Despite the suppression of JA synthesis in tgw1 mutant, ABA synthesis is upregulated, which is accompanied by enhanced expression of SAG3 and reduction of chlorophyll content in leaves. Together, these results demonstrate a role of the JA synthetic gene KAT-2B in controlling GW and its potential application value for wheat improvement.

Emergent Coordination of the CHKB and CPT1B Genes in Eutherian Mammals: Implications for the Origin of Brown Adipose Tissue.

Mitochondrial fatty acid oxidation (FAO) contributes to the proton motive force that drives ATP synthesis in many mammalian tissues. In eutherian (placental) mammals, brown adipose tissue (BAT) can also dissipate this proton gradient through uncoupling protein 1 (UCP1) to generate heat, but the evolutionary events underlying the emergence of BAT are unknown. An essential step in FAO is the transport of cytoplasmic long chain acyl-coenzyme A (acyl-CoA) into the mitochondrial matrix, which requires the action of carnitine palmitoyltransferase 1B (CPT1B) in striated muscle and BAT. In eutherians, the CPT1B gene is closely linked to the choline kinase beta (CHKB) gene, which is transcribed from the same DNA strand and terminates just upstream of CPT1B. CHKB is a rate-limiting enzyme in the synthesis of phosphatidylcholine (PC), a predominant mitochondrial membrane phospholipid, suggesting that the coordinated expression of CHKB and CPT1B may cooperatively enhance mitochondrial FAO. The present findings show that transcription of the eutherian CHKB and CPT1B genes is linked within a unitary epigenetic domain targeted to the CHKB gene, and that that this regulatory linkage appears to have resulted from an intergenic deletion in eutherians that significantly altered the distribution of CHKB and CPT1B expression. Informed by the timing of this event relative to the emergence of BAT, the phylogeny of CHKB-CPT1B synteny, and the insufficiency of UCP1 to account for eutherian BAT, these data support a mechanism for the emergence of BAT based on the acquisition of a novel capacity for adipocyte FAO in a background of extant UCP1.

Effects of fasting, feeding and exercise on plasma acylcarnitines among subjects with CPT2D, VLCADD and LCHADD/TFPD.

BACKGROUND: The plasma acylcarnitine profile is frequently used as a biochemical assessment for follow-up in diagnosed patients with fatty acid oxidation disorders (FAODs). Disease specific acylcarnitine species are elevated during metabolic decompensation but there is clinical and biochemical heterogeneity among patients and limited data on the utility of an acylcarnitine profile for routine clinical monitoring. METHODS: We evaluated plasma acylcarnitine profiles from 30 diagnosed patients with long-chain FAODs (carnitine palmitoyltransferase-2 (CPT2), very long-chain acyl-CoA dehydrogenase (VLCAD), and long-chain 3-hydroxy acyl-CoA dehydrogenase or mitochondrial trifunctional protein (LCHAD/TFP) deficiencies) collected after an overnight fast, after feeding a controlled low-fat diet, and before and after moderate exercise. Our purpose was to describe the variability in this biomarker and how various physiologic states effect the acylcarnitine concentrations in circulation. RESULTS: Disease specific acylcarnitine species were higher after an overnight fast and decreased by approximately 60% two hours after a controlled breakfast meal. Moderate-intensity exercise increased the acylcarnitine species but it varied by diagnosis. When analyzed for a genotype/phenotype correlation, the presence of the common LCHADD mutation (c.1528G > C) was associated with higher levels of 3-hydroxyacylcarnitines than in patients with other mutations. CONCLUSIONS: We found that feeding consistently suppressed and that moderate intensity exercise increased disease specific acylcarnitine species, but the response to exercise was highly variable across subjects and diagnoses. The clinical utility of routine plasma acylcarnitine analysis for outpatient treatment monitoring remains questionable; however, if acylcarnitine profiles are measured in the clinical setting, standardized procedures are required for sample collection to be of value.

The 3-ketoacyl-CoA thiolase: an engineered enzyme for carbon chain elongation of chemical compounds.

Because of their function of catalyzing the rearrangement of the carbon chains, thiolases have attracted increasing attentions over the past decades. The 3-ketoacyl-CoA thiolase (KAT) is a member of the thiolase, which is capable of catalyzing the Claisen condensation reaction between the two acyl-CoAs, thereby achieving carbon chain elongation. In this way, diverse value-added compounds might be synthesized starting from simple small CoA thioesters. However, most KATs are hampered by low stability and poor substrate specificity, which has hindered the development of large-scale biosynthesis. In this review, the common characteristics in the three-dimensional structure of KATs from different sources are summarized. Moreover, structure-guided rational engineering is discussed as a strategy for enhancing the performance of KATs. Finally, we reviewed the metabolic engineering applications of KATs for producing various energy-storage molecules, such as n-butanol, fatty acids, dicarboxylic acids, and polyhydroxyalkanoates. KEY POINTS: • Summarize the structural characteristics and catalyzation mechanisms of KATs. • Review on the rational engineering to enhance the performance of KATs. • Discuss the applications of KATs for producing energy-storage molecules.

Production of polyhydroxyalkanoate from acetate by metabolically engineered Aeromonas hydrophilia.

Aeromonas hydrophila 4AK4 normally produces the copolymer of 3-hydroxybutyrate and 3-hydroxyhexanoate (PHBHHx) using lauric acid as the carbon source. In this study we reported the metabolic engineering of A. hydrophila 4AK4 for the production of polyhydroxyalkanoate (PHA) using acetate as a main carbon source. Recombinant A. hydrophila overexpressing ß-ketothiolase and acetoacetyl-CoA reductase could accumulate poly-3-hydroxybutyrate (PHB) from acetate with a polymer content of 1.39 wt%. Further overexpression of acetate kinase/phosphotransacetylase and acetyl-CoA synthetase improved PHB content to 8.75 wt% and 19.82 wt%, respectively. When acetate and propionate were simultaneously supplied as carbon sources, the engineered A. hydrophila overexpressing ß-ketothiolase, acetoacetyl-CoA reductase, and acetyl-CoA synthetase was found able to produce the copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate (PHBV). The recombinant grew to 3.79 g/L cell dry weight (CDW) containing 15.02 wt% PHBV. Our proposed metabolic engineering strategies illustrate the feasibility for producing PHA from acetate by A. hydrophila.

2-methylacetoacetyl-coenzyme A thiolase (beta-ketothiolase) deficiency: one disease - two pathways.

BACKGROUND: 2-methylacetoacetyl-coenzyme A thiolase deficiency (MATD; deficiency of mitochondrial acetoacetyl-coenzyme A thiolase T2/ "beta-ketothiolase") is an autosomal recessive disorder of ketone body utilization and isoleucine degradation due to mutations in ACAT1. METHODS: We performed a systematic literature search for all available clinical descriptions of patients with MATD. Two hundred forty-four patients were identified and included in this analysis. Clinical course and biochemical data are presented and discussed. RESULTS: For 89.6% of patients at least one acute metabolic decompensation was reported. Age at first symptoms ranged from 2 days to 8 years (median 12 months). More than 82% of patients presented in the first 2 years of life, while manifestation in the neonatal period was the exception (3.4%). 77.0% (157 of 204 patients) of patients showed normal psychomotor development without neurologic abnormalities. CONCLUSION: This comprehensive data analysis provides a systematic overview on all cases with MATD identified in the literature. It demonstrates that MATD is a rather benign disorder with often favourable outcome, when compared with many other organic acidurias.

Transcriptomic Analysis of Glioma Based on IDH Status Identifies ACAA2 as a Prognostic Factor in Lower Grade Glioma.

BACKGROUND: Glioma is the most common and lethal tumor in the central nervous system (CNS). More than 70% of WHO grade II/III gliomas were found to harbor isocitrate dehydrogenase (IDH) mutations which generated targetable metabolic vulnerabilities. Focusing on the metabolic vulnerabilities, some targeted therapies, such as NAMPT, have shown significant effects in preclinical and clinical trials. METHODS: We explored the TCGA as well as CGGA database and analyzed the RNA-seq data of lower grade gliomas (LGG) with the method of weighted correlation network analysis (WGCNA). Differential expressed genes were screened, and coexpression relationships were grouped together by performing average linkage hierarchical clustering on the topological overlap. Clinical data were used to conduct Kaplan-Meier analysis. RESULTS: In this study, we identified ACAA2 as a prognostic factor in IDH mutation lower grade glioma with the method of weighted correlation network analysis (WGCNA). The difference of ACAA2 gene expressions between the IDH wild-type (IDH-WT) group and the IDH mutant (IDH-MUT) group suggested that there may be different potential targeted therapies based on the fatty acid metabolic vulnerabilities, which promoted the personalized treatment for LGG patients.

Primrose syndrome: Characterization of the phenotype in 42 patients.

Primrose syndrome (PS; MIM# 259050) is characterized by intellectual disability (ID), macrocephaly, unusual facial features (frontal bossing, deeply set eyes, down-slanting palpebral fissures), calcified external ears, sparse body hair and distal muscle wasting. The syndrome is caused by de novo heterozygous missense variants in ZBTB20. Most of the 29 published patients are adults as characteristics appear more recognizable with age. We present 13 hitherto unpublished individuals and summarize the clinical and molecular findings in all 42 patients. Several signs and symptoms of PS develop during childhood, but the cardinal features, such as calcification of the external ears, cystic bone lesions, muscle wasting, and contractures typically develop between 10 and 16 years of age. Biochemically, anemia and increased alpha-fetoprotein levels are often present. Two adult males with PS developed a testicular tumor. Although PS should be regarded as a progressive entity, there are no indications that cognition becomes more impaired with age. No obvious genotype-phenotype correlation is present. A subgroup of patients with ZBTB20 variants may be associated with mild, nonspecific ID. Metabolic investigations suggest a disturbed mitochondrial fatty acid oxidation. We suggest a regular surveillance in all adult males with PS until it is clear whether or not there is a truly elevated risk of testicular cancer.

ACAA2 and FASN polymorphisms affect the fatty acid profile of Chios sheep milk.

The objective of the research reported in this research communication was the identification and association of single nucleotide polymorphisms (SNP) in the ovine DGAT1, FASN, SCD1 and ACAA2 genes with milk fat percentage and fatty acid (FA) content. Three consecutive monthly milk samplings were obtained from a total of 429 purebred Chios ewes during mid-lactation. Genotypic data were jointly analyzed with 1184 fat content and 37 718 FA percentage records using mixed models. The 3' untranslated region (UTR) of the DGAT1 gene and the 5' and 3'UTRs of the SCD1 gene appeared to be monomorphic. The FASN g.14777C>T SNP on exon 31 was associated with C13:0 and the ACAA2 g.2982T>C SNP on the 3'UTR was associated with C9:0, C11:0, C12:1 cis-9, C13:0 and the ω6/ω3 index, while fat percentage was not affected by the identified SNPs. The results could be useful for breeding programs aiming to improve the quality and nutritional value of ovine milk.

Oxoglutarate dehydrogenase and acetyl-CoA acyltransferase 2 selectively associate with H2A.Z-occupied promoters and are required for histone modifications.

Histone H2A.Z plays an essential role in regulating transcriptional rates and memory. Interestingly, H2A.Z-bound nucleosomes are located in both transcriptionally active and inactive promotors, with no clear understanding of the mechanisms via which it differentially regulates transcription. We hypothesized that its functions are mediated through recruitment of regulatory proteins to promoters. Using rapid chromatin immunoprecipitation-mass spectrometry, we uncovered the association of H2A.Z-bound chromatin with the metabolic enzymes, oxoglutarate dehydrogenase (OGDH) and acetyl-CoA acyltransferase 2 (ACAA2). Recombinant green florescence fusion proteins, combined with mutations of predicted nuclear localization signals, confirmed their nuclear localization and chromatin binding. Conclusively, chromatin immunoprecipitation-deep sequencing, confirmed the predominant association of OGDH and ACAA2 with H2A.Z-occupied transcription start sites and enhancers, the former of which we confirmed is conserved in both mouse and human tissue. Furthermore, H2A.Z-deficient human HAP1 cells exhibited reduced chromatin-bound metabolic enzymes, accompanied with reduced posttranslational histone modifications, including acetylation and succinylation. Specifically, knockdown of OGDH diminished H4 succinylation. Thus, the data reveal that select metabolic enzymes are assembled at active, H2A.Z-occupied, promoters, for potential site-directed production of metabolic intermediates that are required for histone modifications.

Increase in omega-6 and decrease in omega-3 polyunsaturated fatty acid oxidation elevates the risk of exudative AMD development in adults with Chinese diet.

Appropriate diet is essential for the regulation of age-related macular degeneration (AMD). In particular the type of dietary polyunsaturated fatty acids (PUFA) and poor antioxidant status including carotenoid levels concomitantly contribute to AMD risk. Build-up of oxidative stress in AMD induces PUFA oxidation, and a mix of lipid oxidation products (LOPs) are generated. However, LOPs are not comprehensively evaluated in AMD. LOPs are considered biomarkers of oxidative stress but also contributes to inflammatory response. In this cross-sectional case-control study, plasma omega-6/omega-3 PUFA ratios and antioxidant status (glutathione, superoxide dismutase and catalase), and plasma and urinary LOPs (41 types) were determined to evaluate its odds-ratio in the risk of developing exudative AMD (n = 99) compared to age-gender-matched healthy controls (n = 198) in adults with Chinese diet. The odds ratio of developing exudative AMD increased with LOPs from omega-6 PUFA and decreased from those of omega-3 PUFA. These observations were associated with a high plasma omega-6/omega-3 PUFA ratio and low carotenoid levels. In short, poor PUFA and antioxidant status increased the production of omega-6 PUFA LOPs such as dihomo-isoprostane and dihomo-isofuran, and lowered omega-3 PUFA LOPs such as neuroprostanes due to the high omega-6/omega-3 PUFA ratios; they were also correlated to the risk of AMD development. These findings indicate the generation of specific LOPs is associated with the development of exudative AMD.

MicroRNA-15a Regulates the Differentiation of Intramuscular Preadipocytes by Targeting ACAA1, ACOX1 and SCP2 in Chickens.

Our previous studies showed that microRNA-15a (miR-15a) was closely related to intramuscular fat (IMF) deposition in chickens; however, its regulatory mechanism remains unclear. Here, we evaluated the expression characteristics of miR-15a and its relationship with the expression of acetyl-CoA acyltransferase 1 (ACAA1), acyl-CoA oxidase 1 (ACOX1) and sterol carrier protein 2 (SCP2) by qPCR analysis in Gushi chicken breast muscle at 6, 14, 22, and 30 weeks old, where we performed transfection tests of miR-15a mimics in intramuscular preadipocytes and verified the target gene of miR-15a in chicken fibroblasts (DF1). The miR-15a expression level at 30 weeks increased 13.5, 4.5, and 2.7-fold compared with the expression levels at 6, 14, and 22 weeks, respectively. After 6 days of induction, miR-15a over-expression significantly promoted intramuscular adipogenic differentiation and increased cholesterol and triglyceride accumulation in adipocytes. Meanwhile, 48 h after transfection with miR-15a mimics, the expression levels of ACAA1, ACOX1 and SCP2 genes decreased by 56.52%, 31.18% and 37.14% at the mRNA level in intramuscular preadipocytes. In addition, the co-transfection of miR-15a mimics and ACAA1, ACOX1 and SCP2 3'UTR (untranslated region) dual-luciferase vector significantly inhibited dual-luciferase activity in DF1 cells. Taken together, our data demonstrate that miR-15a can reduce fatty acid oxidation by targeting ACAA1, ACOX1, and SCP2, which subsequently indirectly promotes the differentiation of chicken intramuscular preadipocytes.

Mutation update on ACAT1 variants associated with mitochondrial acetoacetyl-CoA thiolase (T2) deficiency.

Mitochondrial acetoacetyl-CoA thiolase (T2, encoded by the ACAT1 gene) deficiency is an inherited disorder of ketone body and isoleucine metabolism. It typically manifests with episodic ketoacidosis. The presence of isoleucine-derived metabolites is the key marker for biochemical diagnosis. To date, 105 ACAT1 variants have been reported in 149 T2-deficient patients. The 56 disease-associated missense ACAT1 variants have been mapped onto the crystal structure of T2. Almost all these missense variants concern residues that are completely or partially buried in the T2 structure. Such variants are expected to cause T2 deficiency by having lower in vivo T2 activity because of lower folding efficiency and/or stability. Expression and activity data of 30 disease-associated missense ACAT1 variants have been measured by expressing them in human SV40-transformed fibroblasts. Only two variants (p.Cys126Ser and p.Tyr219His) appear to have equal stability as wild-type. For these variants, which are inactive, the side chains point into the active site. In patients with T2 deficiency, the genotype does not correlate with the clinical phenotype but exerts a considerable effect on the biochemical phenotype. This could be related to variable remaining residual T2 activity in vivo and has important clinical implications concerning disease management and newborn screening.