NSPlex: an efficient method to analyze non-specific peaks amplified using commercial STR kits.

Commercial short tandem repeat (STR) kits exclusively contain human-specific primers; however, various non-human organisms with high homology to the STR kit's primer sequences can cause cross-reactivity. Owing to the proprietary nature of the primers in STR kits, the origins and sequences of most non-specific peaks (NSPs) remain unclear. Such NSPs can complicate data interpretation between the casework and reference samples; thus, we developed "NSPlex", an efficient method to discover the biological origins of NSPs. We used leftover STR kit amplicons after capillary electrophoresis and performed advanced bioinformatics analyses using next-generation sequencing followed by BLAST nucleotide searches. Using our method, we could successfully identify NSP generated from PCR amplicons of a sample mixture of human DNA and DNA extracted from matcha powder (finely ground powder of green tea leaves and previously known as a potential source of NSP). Our results showed our method is efficient for NSP analysis without the need for the primer information as in commercial STR kits.

Unfavorable switching of skewed X chromosome inactivation leads to Menkes disease in a female infant.

Menkes disease is an X-linked disorder of copper metabolism caused by mutations in the ATP7A gene, and female carriers are usually asymptomatic. We describe a 7-month-old female patient with severe intellectual disability, epilepsy, and low levels of serum copper and ceruloplasmin. While heterozygous deletion of exons 16 and 17 of the ATP7A gene was detected in the proband, her mother, and her grandmother, only the proband suffered from Menkes disease clinically. Intriguingly, X chromosome inactivation (XCI) analysis demonstrated that the grandmother and the mother showed skewing of XCI toward the allele with the ATP7A deletion and that the proband had extremely skewed XCI toward the normal allele, resulting in exclusive expression of the pathogenic ATP7A mRNA transcripts. Expression bias analysis and recombination mapping of the X chromosome by the combination of whole genome and RNA sequencing demonstrated that meiotic recombination occurred at Xp21-p22 and Xq26-q28. Assuming that a genetic factor on the X chromosome enhanced or suppressed XCI of its allele, the factor must be on either of the two distal regions derived from her grandfather. Although we were unable to fully uncover the molecular mechanism, we concluded that unfavorable switching of skewed XCI caused Menkes disease in the proband.

Lack of GPR180 ameliorates hepatic lipid depot via downregulation of mTORC1 signaling.

Our previous genome-wide association study to explore genetic loci associated with lean nonalcoholic fatty liver disease (NAFLD) in Japan suggested four candidate loci, which were mapped to chr6, chr7, chr12 and chr13. The present study aimed to identify the locus involved functionally in NAFLD around the association signal observed in chr13. Chromosome conformation capture assay and a database survey suggested the intermolecular interaction among DNA fragments in association signals with the adjacent four coding gene promoters. The four genes were further screened by knockdown (KD) in mice using shRNA delivered by an adeno-associated virus vector (AAV8), and KD of G protein-coupled receptor 180 (Gpr180) showed amelioration of hepatic lipid storage. Gpr180 knockout (KO) mice also showed ameliorated hepatic and plasma lipid levels without influencing glucose metabolism after high-fat diet intake. Transcriptome analyses showed downregulation of mTORC1 signaling and cholesterol homeostasis, which was confirmed by weakened phosphorylation of mTOR and decreased activated SREBP1 in Gpr180KO mice and a human hepatoma cell line (Huh7). AAV8-mediated hepatic rescue of GPR180 expression in KO mice showed recovery of plasma and hepatic lipid levels. In conclusion, ablation of GPR180 ameliorated plasma and hepatic lipid levels, which was mediated by downregulation of mTORC1 signaling.

KBTBD11, encoding a novel PPARγ target gene, is involved in NFATc1 proteolysis by interacting with HSC70 and HSP60.

We previously revealed that Kbtbd11 mRNA levels increase during 3T3-L1 differentiation and Kbtbd11 knockdown suppresses whereas its overexpression promotes adipogenesis. However, how Kbtbd11 mRNA is regulated during adipocyte differentiation and how the KBTBD11 protein functions in adipocytes remain elusive. This study aimed to examine the transcriptional regulatory mechanism of Kbtbd11 during adipocyte differentiation, KBTBD11-interacting protein functions, and elucidate the role of KBTBD11 in adipocytes. First, we identified the PPRE consensus sequences in the Kbtbd11 exon 1- and intron 1-containing region and demonstrated that PPARγ acts on this region to regulate Kbtbd11 expression. Next, we purified the KBTBD11 protein complex from 3T3-L1 adipocytes and identified heat shock proteins HSC70 and HSP60 as novel KBTBD11-interacting proteins. HSC70 and HSP60 inhibition increased KBTBD11 protein levels that promoted NFATc1 ubiquitination. These data suggest that HSC70 and HSP60 are involved in KBTBD11 stabilization and are responsible for NFATc1 regulation on the protein level. In summary, this study describes first the protein regulatory mechanism of NFATc1 through the HSC70/HSP60-KBTBD11 interaction that could provide a potential new target for the differentiation and proliferation of various cells, including adipocytes and tumors.

Probiotics suppress nonalcoholic steatohepatitis and carcinogenesis progression in hepatocyte-specific PTEN knockout mice.

Nonalcoholic fatty liver disease (NAFLD), a hepatic characteristic of metabolic syndrome, received significant attention in clinical settings. The multiple-hit theory is one of the proposed mechanisms of NAFLD, and gut dysbiosis is considered a hit. Thus, controlling gut microbiota is a potential target in the management of NAFLD, and probiotics can be used as a treatment agent for NAFLD. The current study aimed to investigate the efficacy of probiotics against nonalcoholic steatohepatitis in a hepatocyte-specific PTEN knockout mouse model that mimics the characteristics of human NAFLD. Probiotics were administered to male knockout mice for 8 or 40 weeks. Next, we assessed hepatic inflammation, fibrosis, carcinogenesis, and oxidative stress. Probiotics were found to reduce serum transaminase levels, NAFLD activity score, and the gene expression of pro-inflammatory cytokines. In addition, they decreased liver fibrosis grade, which was examined via Sirius red staining, gene expression of fibrotic markers, and hydroxyproline. Furthermore, probiotics suppressed the number of liver tumors, particular in HCC. Probiotics reduced oxidative stresses, including glutathione levels, and anti-oxidative stress marker, which may be an underlying mechanism for their beneficial effects. In conclusion, probiotics treatment had beneficial effects against NAFLD and carcinogenesis in hepatocyte-specific PTEN knockout mice.

Genome-wide DNA methylation status of Mongolians exhibits signs of cellular stress response related to their nomadic lifestyle.

BACKGROUND: Epigenetics is crucial for connecting environmental stresses with physiological responses in humans. Mongolia, where nomadic livestock pastoralism has been the primal livelihood, has a higher prevalence of various chronic diseases than the surrounding East Asian regions, which are more suitable for crop farming. The genes related to dietary stress and pathogenesis of related disorders may have varying epigenetic statuses among the human populations with diverse dietary cultures. Hence, to understand such epigenetic differences, we conducted a comparative analysis of genome-wide DNA methylation of Mongolians and crop-farming East Asians. METHODS: Genome-wide DNA methylation status of peripheral blood cells (PBCs) from 23 Mongolian adults and 24 Thai adults was determined using the Infinium Human Methylation 450K arrays and analyzed in combination with previously published 450K data of 20 Japanese and 8 Chinese adults. CpG sites/regions differentially methylated between Mongolians and crop-farming East Asians were detected using a linear model adjusted for sex, age, ethnicity, and immune cell heterogeneity on RnBeads software. RESULTS: Of the quality-controlled 389,454 autosomal CpG sites, 223 CpG sites were significantly differentially methylated among Mongolians and the four crop farming East Asian populations (false discovery rate < 0.05). Analyses focused on gene promoter regions revealed that PM20D1 (peptidase M20 domain containing 1), which is involved in mitochondrial uncoupling and various processes, including cellular protection from reactive oxygen species (ROS) and thermogenesis, was the top differentially methylated gene. Moreover, gene ontology enrichment analysis revealed that biological processes related to ROS metabolism were overrepresented among the top 1% differentially methylated genes. The promoter regions of these genes were generally hypermethylated in Mongolians, suggesting that the metabolic pathway detoxifying ROS might be globally suppressed in Mongolians, resulting in the high susceptibility of this population to various chronic diseases. CONCLUSIONS: This study showed a significantly diverse DNA methylation status among Mongolians and crop-farming East Asians. Further, we found an association between the differentially methylated genes and various metabolic and neurodegenerative diseases. Knowledge of the epigenetic regulators might help in proper understanding, treatment, and control of such disorders, and physiological adaptation in the future.

A case of congenital fiber-type disproportion syndrome presenting dilated cardiomyopathy with ACTA1 mutation.

BACKGROUND: Actin, alpha, skeletal muscle 1 (ACTA1) is one of the causative genes of nemaline myopathy (NM) and congenital fiber-type disproportion (CFTD). CFTD is characterized by type 1 fiber atrophy and distinguished from NM in the absence of rods. Eight patients with CFTD, including one patient with dilated cardiomyopathy (DCM), have previously been reported. Herein, we report the case of a 10-year-old boy presenting with CFTD and DCM. METHODS: We performed exome sequencing and analyzed the effect of Met327Lys mutations on cultured C2C12 muscle cells compared with that seen in the wild type (WT, ACTA1) and previously identified Asp294Val mutations associated with a severe phenotype of CFTD without cardiomyopathy. RESULTS: Exome sequencing revealed a de novo mutation, c.980 T > A, p.(Met327Lys), in ACTA1 (NM_001100.4). C2C12 cells transfected with the WT plasmid expressed ACTA1 in the nucleus and cytoplasm. Cells with the Asp294Val mutant showed needle-like structures in the cytoplasm, whereas the expression of the Met327Lys mutant resulted in few aggregations but many apoptotic cells. CONCLUSION: Apoptosis induced in Met327Lys-transfected muscle cells supports the pathogenicity of the mutation and can be implicated as one of the histopathological features associated with CFTD, as in NM.

Investigation of Maternal Diet and FADS1 Polymorphism Associated with Long-Chain Polyunsaturated Fatty Acid Compositions in Human Milk.

Increasing the amount of long-chain polyunsaturated fatty acids (LCPUFA) in human milk is an important strategy for infant growth and development. We investigated the associations of LCPUFA compositions in human milk with maternal diet (especially fish and shellfish intake), with fatty acid Δ5 desaturase gene (FADS1) polymorphisms, and with gene-diet interactions. The present study was performed as part of an adjunct study of the Japan Environment and Children's Study. The participants were 304 lactating females, who provided human milk 6−7 months after delivery. Fatty acids in human milk were analyzed by gas chromatography, and dietary surveys were conducted using a brief self-administered diet history questionnaire. We also analyzed a single nucleotide polymorphism of FADS1 (rs174547, T/C). There was a significant difference in arachidonic acid (ARA) composition in human milk among the genotype groups, and the values were decreasing in the order of TT > TC > CC. The concentrations of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were also different between TT and CC genotype, indicating a tendency for decreasing values in the same order. The composition of ARA showed significant gene−dietary interactions in multiple regression analysis, and the positive correlation between fish and shellfish intake and ARA composition in human milk was significant only in the CC genotype. Moreover, the factor most strongly associated with EPA and DHA composition in human milk was fish and shellfish intake. Therefore, it was suggested that increasing fish and shellfish intake in mothers may increase EPA and DHA composition in human milk, while increasing fish and shellfish intake in CC genotype mothers may lead to increased ARA composition in human milk.

Association of HLA-DPB1, NLRP10, OVOL1, and ABCC11 with the axillary microbiome in a Japanese population.

BACKGROUND: The distinct diversity of the human skin microbiome depends not only on the body site but also the individual. Host-commensal interactions have been described for the gut microbiome, but little is known about the epidermal microbiome. OBJECTIVE: The present study investigated whether genetic variants associated with skin traits affect the axillary microbiome. METHODS: Eight skin trait-related single nucleotide polymorphisms and HLA-A, -B, -C, and -DPB1 were genotyped in 186 Japanese males. From axillary swabs, the intensity of a representative axillary odor, trans (E) isomer of 3-methyl-2-hexenoic acid (E3M2H), was quantified with gas chromatography-tandem mass spectrometry analysis, the diversity of the axillary microbiome was evaluated with a 16 s rRNA metagenomic approach, and the association of these characteristics was assessed statistically. RESULTS: A risk allele for atopic dermatitis of rs878860 in NLRP10 and the allele for wet earwax of rs17822931 in ABCC11 decreased the relative abundance of Corynebacterium. Conversely, these alleles increased the relative abundance of Staphylococcus. Metagenomic analysis revealed that ß-diversity showed significant dissimilarity at the weighted Unifrac distance between minor allele carrier and non-carrier groups in HLA-DPB1*05:01, rs17822931, and rs878860. HLA-DPB1*04:01, HLA-DPB1*05:01, and rs17822931 were associated with E3M2H. CONCLUSIONS: We identified novel candidate loci associated with the axillary microbiome and malodor.

N4BP2L1 interacts with dynactin and contributes to GLUT4 trafficking and glucose uptake in adipocytes.

AIMS/INTRODUCTION: It was reported previously that N4bp2l1 expression increases in 3T3-L1 cells in a differentiation-dependent manner and N4bp2l1 knockdown suppresses adipocyte differentiation. However, the physiological function of N4BP2L1 in adipocytes remains unknown. This study aimed to elucidate the physiological mechanism of N4bp2l1 expression and the role of N4BP2L1 in the physiological function of adipocytes. MATERIALS AND METHODS: Analysis of gene expression levels of N4bp2l1 in adipose tissue during feeding in mice was conducted. Identification of transcription factors that regulate N4bp2l1 expression was conducted using a reporter assay. Investigation of N4BP2L1-interacting proteins was carried out using immunoprecipitation. A GLUT4 translocation assay and a glucose uptake assay in 3T3-L1 adipocytes were performed using N4bp2l1 overexpression and knockdown adenovirus. RESULTS: The results indicated that N4bp2l1 is a novel FoxO1 target gene and its expression is controlled by the insulin-mediated regulation of FoxO1. N4BP2L1 interacts with dynactin, which binds to the microtubule motor dynein, indicating that N4BP2L1 is involved in GLUT4 trafficking and glucose uptake in 3T3-L1 adipocytes. CONCLUSIONS: Our results suggest that N4BP2L1 is involved in adipocyte homeostasis by interacting with dynein-dynactin and affecting GLUT4-mediated glucose uptake and the insulin signaling pathway.

Two cases of DYNC1H1 mutations with intractable epilepsy.

BACKGROUND: The DYNC1H1 gene encodes the heavy chain of cytoplasmic dynein 1, a core structure of the cytoplasmic dynein complex. Dominant DYNC1H1 mutations are implicated in Charcot-Marie-Tooth disease, axonal, type 20, spinal muscular atrophy, lower extremity-predominant 1, and autosomal dominant mental retardation 13 with neuronal migration defects. We report two patients with DYNC1H1 mutations who had intractable epilepsy and intellectual disability (ID), one with and one without pachygyria. CASE REPORTS: Patient 1 had severe ID. At the age of 2 months, she presented myoclonic seizures and tonic seizures, and later experienced atonic seizures and focal impaired-awareness seizures (FIAS). EEG showed slow waves in right central areas during myoclonic seizures. Brain MRI revealed pachygyria, predominantly in the occipital lobe. After callosal transection her atonic seizures disappeared, but FIAS remained. Patient 2 was diagnosed with autism spectrum disorder (ASD) and severe ID. At the age of 7 years, he presented generalized tonic-clonic seizures, myoclonic seizures, and FIAS. Interictal EEG showed generalized spike-and-wave complexes, predominantly in the left frontal area. Brain MRI was unremarkable. Exome sequencing revealed novel de novo mutations in DYNC1H1: c.4691A > T, p.(Glu1564Val) in Patient 1 and c.12536 T > C, p.(Leu4179Ser) in Patient 2. CONCLUSIONS: DYNC1H1 comprises a stem, stalk, and six AAA domains. Patient 2 is the second report of an AAA6 domain mutation without malformations of cortical development. The p.(Gly4072Ser) mutation in the AAA6 domain was also reported in a patient with ASD. It may be that the AAA6 domain has little effect on neuronal movement of DYNC1H1 along microtubules.

Normal plasma apoB48 despite the virtual absence of apoB100 in a compound heterozygote with novel mutations in the MTTP gene.

"Normotriglyceridemic abetalipoproteinemia (ABL)" was originally described as a clinical entity distinct from either ABL or hypobetalipoproteinemia. Subsequent studies identified mutations in APOB gene which encoded truncated apoB longer than apoB48. Therefore, "Normotriglyceridemic ABL" can be a subtype of homozygous familial hypobetalipoproteinemia. Here, we report an atypical female case of ABL who was initially diagnosed with "normotriglyceridemic ABL", because she had normal plasma apoB48 despite the virtual absence of apoB100 and low plasma TG level. Next generation sequencing revealed that she was a compound heterozygote of two novel MTTP mutations: nonsense (p.Q272X) and missense (p.G709R). We speculate that p.G709R might confer residual triglyceride transfer activity of MTTP preferentially in the intestinal epithelium to the hepatocytes, allowing production of apoB48. Together, "normotriglyceridemic ABL" may be a heterogenous disorder which is caused by specific mutations in either APOB or MTTP gene.

ILDR2 stabilization is regulated by its interaction with GRP78.

Ildr2 was initially identified as a genetic modifier of diabetes susceptibility in B6.DBA Lepob congenic mice, and was associated with decreased ß-cell replication rates, reduced ß-cell mass, and persistent mild hypoinsulinemic hyperglycemia. However, the molecular mechanisms of how the ILDR2 protein is involved in these effects are largely unknown. We sought to identify ILDR2-interacting proteins to further elucidate the molecular mechanisms underpinning ILDR2 function in pancreatic ß-cells. Using TAP tag technology, we purified proteins interacting with ILDR2 in the pancreatic ß-cell line MIN6, and identified the endoplasmic reticulum resident chaperones, GRP78 and PDIA1, as novel proteins interacting with ILDR2. We demonstrated that GRP78 interacted with ILDR2 and was possibly involved in ILDR2 stabilization by inhibiting ubiquitin-proteasome degradation. Additionally, adenoviral ILDR2 knockdown led to reduced glucose-responsive insulin secretion in MIN6 ß-cells, suggesting ILDR2 may be implicated in a new pathway in hypoinsulinemic hyperglycemia. These data provide evidence for a novel association between GRP78 and ILDR2, and suggest GPR78-ILDR2 may a novel target for diabetic therapeutic modulation in decreased insulin secretion.

Novel BEST1 mutation in autosomal recessive bestrophinopathy in Japanese siblings.

PURPOSE: Autosomal recessive bestrophinopathy (ARB) is a disease that results from the mutations in the BEST1 gene. It is characterized by multifocal yellowish lipofuscin deposits, cystoid macular edema, and subretinal fluid. Among approximately 270 BEST1 mutations, only 40 that include both heterozygous and homozygous mutations are associated with ARB. However, very few ARB-related mutations have been reported in the Japanese population. Therefore, in this study, we aimed to identify BEST1 mutations and describe the genotype-phenotype relationship in Japanese dizygotic twins presenting with ARB. MATERIALS AND METHODS: We performed clinical examinations in Japanese dizygotic twin patients (male: 29 years) with ARB as well as whole-exome sequencing in seven family members of these twins. RESULTS: In this study, we have reported on a novel BEST1 mutation, the p. Phe151Cys mutation, associated with ARB in Japanese dizygotic twins who had bi-allelic p. Ala160Pro mutations in BEST1. The clinical features observed were binocular abnormalities of the fundus, such as multifocal yellowish subretinal deposits, cystoid macular edema, and subretinal fluid. The full-field electroretinography results were subnormal. CONCLUSION: It was indicated that the novel BEST1 mutations identified may be strongly correlated with binocular ARB. This study provides significant information of the genotype-phenotype association in Japanese ARB patients. Further, the genetic analysis that we performed was very useful for the differential diagnosis and might have implications in the development of future treatment modalities.

Genome-Wide Association Study of Lean Nonalcoholic Fatty Liver Disease Suggests Human Leukocyte Antigen as a Novel Candidate Locus.

Nonalcoholic fatty liver disease (NAFLD) is supposed to manifest its metabolic phenotype in the liver, but it is common to have lean individuals diagnosed with NAFLD, known as lean NAFLD. We conducted a two-stage analysis to identify NAFLD-associated loci in Japanese patients. In stage I, 275 metabolically healthy normal-weight patients with NAFLD were compared with 1,411 non-NAFLD controls adjusted for age, sex, and alcohol consumption by a genome-wide association study (GWAS). In stage II, human leukocyte antigen (HLA) in chromosome 6 (chr6) (P = 6.73E-08), microRNA (MIR) MIR548F3 in chr7 (P = 4.25E-07), myosin light chain 2 (MYL2) in chr12 (P = 4.39E-07), and glycoprotein precursor (GPC)6 in chr13 (P = 5.43E-07), as suggested by the GWAS, were assessed by single nucleotide polymorphism (SNP) association analysis of whole NAFLD against non-NAFLD in 9,726 members of the general population. A minor allele of the secondary lead SNP in chr6, rs2076529, was significantly associated (odds ratio [OR], 1.19; 95% confidence interval [CI], 1.11-1.28; P = 2.10E-06) and the lead SNP in chr7 was weakly associated (OR 1.15; 95% CI, 1.04-1.27; P = 6.19E-03) with increased NAFLD risk. Imputation-based typing of HLA showed a significant difference in the distribution of HLA-B, HLA-DR-beta chain 1 (DRB1), and HLA-DQ-beta chain 1 (DQB1) alleles in lean NAFLD GWAS. Next-generation sequence-based typing of HLA in 5,649 members of the general population replicated the significant difference of HLA-B allele distribution and the significant increase of the HLA-B*54:01 allele in whole NAFLD. Fecal metagenomic analysis of 3,420 members of the general population showed significant dissimilarity in beta-diversity analysis of rs2076529 and HLA-B*54:01 allele carriers from noncarriers. Veillonellaceae was increased but Verrucomicrobia was decreased in rs2076529 minor allele and HLA-B*54:01 allele carriers as in NAFLD. Conclusion: HLA was identified as a novel locus associated with NAFLD susceptibility, which might be affected by the alteration of gut microbiota.

Serum haptoglobin correlates positively with cholesterol and triglyceride concentrations in an obese Mongolian population.

BACKGROUND: Recent studies revealed that several genetic polymorphisms of haptoglobin gene (HP) and the haptoglobin-related protein gene (HPR) associated not only with haptoglobin (HP) but total, non-HDL, and/or LDL cholesterol concentrations in various populations. METHODS: Association between serum HP concentrations and polymorphisms of HP and the HPR gene, or anthropometric and metabolic factors were examined in Mongolian participants (n = 927) using linear regression analyses. RESULTS: The association of HP and HPR polymorphisms with serum HP concentration but not serum lipids concentrations was observed. However, subgroup analysis revealed that the association of HP and HPR polymorphisms with serum HP concentration was weakened in subgroup of obese (BMI ≥ 30) subjects and positive correlations between serum HP and non-HDL cholesterol, HDL cholesterol or triglyceride concentrations were observed in the obese subjects as compared with in subgroups of normal weight (BMI < 25) and overweight (25 ≤ BMI < 30) subjects. CONCLUSION: The degree of obesity strongly affects the relationships between serum HP concentrations and several genetic, anthropometric and metabolic factors. These results suggested that we need to take into account the degree of obesity when considering the HP polymorphisms as predictive markers for clinical states.

Associations of erythrocyte fatty acid compositions with FADS1 gene polymorphism in Japanese mothers and infants.

Long-chain polyunsaturated fatty acids (LC-PUFAs) are involved in the fetal growth in utero, and are essential for the development of visual and cognitive functions during infancy. The purpose of this study was to examine the associations of erythrocyte fatty acid compositions with FADS1 gene polymorphism in Japanese mothers and infants. The subjects were 383 mothers who participated in an adjunct birth cohort study of the Japan Environment and Children's Study (JECS). In maternal FADS1 SNP genotypes, the precursor fatty acids composition of the Δ5 desaturase in the maternal blood showed significant differences in levels among the groups, and showed increasing values in the order of TT < TC < CC genotype groups. On the other hand, many product fatty acids levels were significantly reduced in the order of TT > TC > CC genotype groups, and DHA levels were significantly lower in the CC genotype group relative to the other groups. Likewise, the relationship between fetal genotype group and fatty acid composition in cord blood was very similar to the maternal relationship. These results indicate the maternal and fetal blood fatty acid compositions are strongly influenced by the FADS1 genotypes. With respect to the cord blood DHA composition, the levels in the fetal CC genotype group showed a trend toward lower values in the maternal CC genotype group pair (p = 0.066) compared to the maternal TC genotype group pair. However, in the fetal TT and TC genotype groups (p = 0.131, p = 0.729, respectively), the maternal genotype did not have a significant effect. The DHA composition was more influenced by the maternal genotype in the fetal CC genotype group than in the fetal TT and TC genotype groups. It was shown that DHA transport via the placenta from the mother might be promoted in the fetal CC genotype compared to the other fetal genotype groups. In conclusion, differences in the FADS1 SNP genotypes of pregnant women and their children may greatly affect the supply of LC-PUFAs. Further studies on the involvement of the FADS1 polymorphisms and the fetal LC-PUFA levels in the fetal growth and development are warranted.

Kbtbd11 contributes to adipocyte homeostasis through the activation of upstream stimulatory factor 1.

The present study aimed to investigate the transcriptional regulation of Kbtbd11 in adipose tissue. To elucidate the physiological role of Kbtbd11 gene expression, adipose Kbtbd11 mRNA expression levels were estimated under various feeding states in wild-type mice. Kbtbd11 expression increased in a time-dependent manner in the adipose tissue in mice fed on chow diet, whereas the promotion of Kbtbd11 mRNA expression by refeeding was attenuated in mice fed on high-fat (HF) diet, suggesting the suppression of Kbtbd11 mRNA expression under HF diets and that changes in mRNA levels were associated with regulation of the transcription activity of Kbtbd11 by some transcription factors. To investigate the transcriptional regulation of Kbtbd11, the fragment upstream of either mouse Kbtbd11 or human KBTBD11 promoter was inserted into a luciferase vector. Luciferase reporter assays revealed that both mouse and human KBTBD11 promoter activity was increased by USF1. Direct USF1 binding to the Ebox in the Kbtbd11 promoter was confirmed by electrophoretic mobility shift and chromatin immunoprecipitation assays. In addition, the adipocyte differentiation marker levels increased instantly in Kbtbd11-overexpressing Usf1 knockdown cells than in Usf1 knockdown cells. These results imply an association of between Kbtbd11 with Usf1 expression and suggest the involvement of Kbtbd11 in a novel adipogenesis pathway.

A novel upstream transcription factor 1 target gene N4bp2l1 that regulates adipogenesis.

N4BP2l1, which is highly expressed in oral squamous cell carcinoma, is associated with poor prognosis. However, N4bp2l1's role in adipogenesis remains unknown. We aimed to clarify the expression profile and transcriptional regulation of N4bp2l1 to elucidate the functions underlying the role of N4bp2l1 in adipocyte differentiation. Our results revealed that N4bp2l1 mRNA expression increased in 3T3-L1 cells in a differentiation-dependent manner. To investigate the transcriptional regulation of N4bp2l1, the 2-kb 5' region upstream of the mouse N4bp2l1 promoter was cloned into a luciferase vector. Luciferase reporter assays indicated that USF1 induces the N4bp2l1 promoter activity. Electrophoretic mobility shift and chromatin immunoprecipitation assays confirmed that USF1 directly binds to the Ebox in the N4bp2l1 promoter. Furthermore, the expressions of adipocyte differentiation markers significantly decreased in N4bp2l1-knockdown cells compared with those in control cells. Our results demonstrated that N4bp2l1 is a novel USF1 target gene that may be involved in adipogenesis regulation.