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Background We previously showed that patients without pathogenic variants in the LDLR and PCSK9 genes comprised approximately 40% of familial hypercholesterolemia (FH) cases. Objective Our aim was to identify novel causative variants in Japanese patients with FH. Methods Whole-exome sequencing was performed in 216 family members from 123 families without pathogenic variants in the LDLR and PCSK9 genes. Clinical and biochemical data were gathered from the family members. Results The known p.(Arg3527Gln) variant in the APOB gene was identified in one Japanese family. The other pathogenic variants in the APOB gene were not identified. The p.(Arg3527Gln) variant was not identified in the other 113 index cases without pathogenic variants in the LDLR and PCSK9 genes. The allele frequency of the p.(Arg3527Gln) variant was 0.0001 in the general Japanese population. Conclusion This is the first report of Japanese cases of FH caused by a known pathogenic APOB variant, p.(Arg3527Gln).

Genes can influence lipid profile and anthropometric variables related to obesity. The present study aimed to verify if variants of the APOE, APOB, ADIPOQ, HSD11β1, and PLIN4 genes are associated with lipid levels or anthropometric variables in a sample comprised of 393 Euro-Brazilian children and adolescents. DNA was genotyped by TaqMan allelic discrimination assay. The ε4 and ε2 alleles of the APOE gene were associated respectively with lower high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) levels (p=0.015 and p=0.012, respectively), while the ε3 allele was associated with higher abdominal circumference (p=0.0416) and excess weight (p=0.0001). The G allele (rs846910) of the HSD11β1 gene was also associated with excess weight (p=0.039). No other association was found. Our results indicate that the ε4 and ε2 alleles could contribute to lower HDL-C and LDL-C levels, respectively, furthermore, the ε3 allele and the G allele (rs846910) of HSD11β1 gene may be risk factors for excess of weight.These findings are very important because we observed that some genetic variants influence the lipid profile and anthropometric variables early in life.

Apolipoprotein B (ApoB) is a major protein component of plasma lipoproteins. It is involved in many important biological processes such as lipid transportation, enzyme activity regulation, and receptor recognition. Extensive studies have shown that the expression of ApoB is regulated at multiple levels. However, the regulation of ApoB expression by microRNAs (miRNAs) still remains unknown. In the present study, identified are miRNAs that are predicted to interact with ApoB in chicken. The predicted relationship between the identified miRNAs and ApoB was verified through dual luciferase reporter assay in chicken DF1 cells, and the effect of miRNAs on ApoB expression was analyzed in chicken embryo hepatocytes stimulated by 17β-estradiol. The results show that miR-101-2-5p was predicted to interact with ApoB. Dual luciferase reporter assay together with the miR-101-2-5p mimics study demostrate that ApoB is the target of miR-101-2-5p, which suppresses the expression of ApoB through binding with the 3′UTR of ApoB. Our experiments suggest that miR-101-2-5p might be involved in lipid metabolism through binding to the 3′UTR of ApoB in the liver of egg-laying chickens.

Familial hypercholesterolemia (FH) is a monogenic dominant inherited disorder of lipid metabolism characterized by elevated low-density lipoprotein levels, and is mainly attributable to mutations in low-density lipoprotein receptor (LDLR), apolipoprotein B (APOB), and proportein convertase subtilisin/kexin type 9 (PCSK9) genes. Next-generation and exome sequencing studies have primarily involved genome-wide association analyses, and meta-analyses and next-generation studies examined a few single-nucleotide polymorphisms (rs151009667 and Val2095Glu) in the ApoB gene. The present study was conducted to investigate the association of APOB and patients with FH in a Saudi population. We genotyped 100 patients with FH and 100 controls for 2 polymorphisms in APOB using polymerase chain reaction-restriction fragment length polymorphism, followed by 3% agarose gel electrophoresis. The strength of the association between the genotype and allele frequencies with the risk of developing FH was evaluated. Clinical details and genotype analysis results were recorded. For the rs151009667 polymorphism, 18% of the CT genotypes were observed only in patients with FH. There was a positive association between CT and CC (odds ratio [OR] 45.07 [95% conflict of interest (CI), 2.67–759.1]; P = .0001) and between T and C (OR 87.8 [95% CI, 5.34–144.2]; P

Background: Familial hypobetalipoproteinemia type 1 (FHBL-1) is a codominant disorder characterized by greatly reduced plasma levels of total cholesterol, low-density lipoprotein cholesterol, and apolipoprotein B. Rare exonic pathogenic variants of APOB gene (nonsense variants, minute deletions/insertions and nonsynonymous variants) have been frequently reported in subjects with FHBL-1. Also, rare intronic variants of APOB located at intron/exon junctions and assumed to affect splicing have been reported. However, the pathogenicity of most of these intronic variants remains to be established. Objective: The objective of this study was the in vitro functional characterization of six splicing variants of APOB gene identified in seven putative FHBL-1 heterozygotes. Methods: ApoB minigenes harboring each variant were expressed in COS-1 cells and their transcripts were sequenced. Results: Four novel variants (c.237+1G>A, c.818+5G>A, c.3000-1G>T, and c.3842+1G>A), predicted in silico to obliterate splice site activity, were found to generate abnormal transcripts. The abnormal transcripts were generated by the activation of cryptic splice sites or exon skipping. All these transcripts harbored a premature termination codon and were predicted to encode truncated apoBs devoid of function. The predicted translation products were: i) p.(Lys41Serfs*2) and p.(Val80Ilefs*10) for c.237+1G>A; ii) p.(Asn274*) for c.818+5G>A; iii) p.(Leu1001Alafs*10) for c.3000-1G>T, and iv) p.(Ser1281Argfs*2) for c.3842+1G>A. Two previously annotated rare variants (c.905-15C>G and c.1618-4G>A) with uncertain effect in silico were found to generate only wild-type transcripts. Conclusions: These in vitro minigene expression studies support the assignment of pathogenicity to four novel splice site variants of APOB gene found in FHBL-1.

Oxidatively modified low-density lipoproteins (oxLDL) play an important role in the occurrence and progression of atherosclerosis. To identify the genetic factors influencing the oxLDL levels, we have genotyped 776 DNA samples of Russian individuals for 196,725 single-nucleotide polymorphisms (SNPs) using the Cardio-MetaboChip (Illumina, USA) and conducted genome-wide association study (GWAS). Fourteen common variants in the locus including APOB gene were significantly associated with the oxLDL levels (P < 2.18 × 10-7). These variants explained only 6% of the variation in the oxLDL levels. Then, we assessed the contribution of rare coding variants of APOB gene to the oxLDL levels. Individuals with the extreme oxLDL levels (48 with the lowest and 48 with the highest values) were selected for targeted sequencing of the region including APOB gene. To evaluate the contribution of the SNPs to the oxLDL levels we used various statistical methods for the association analysis of rare variants: WST, SKAT, and SKAT-O. We revealed that both synonymous and nonsynonymous SNPs affected the oxLDL levels. For the joint analysis of the rare and common variants, we conducted the SKAT-C testing and found a group of 15 SNPs significantly associated with the oxLDL levels (P = 2.14 × 10-9). Our results indicate that the oxLDL levels depend on both common and rare variants of the APOB gene.

Purpose of Review Familial hypercholesterolaemia (FH) is an inherited disorder of low-density lipoprotein cholesterol (LDL-C) which is characterised by a raised cholesterol level from birth and a high risk of premature coronary heart disease. In this paper, we review the genetic basis of FH and its impact on the clinical presentation. Recent Findings Mutations in any of three genes (LDLR, APOB and PCSK9) are known to cause autosomal dominant FH, but a mutation can be found in only ∼40% of patients with a clinical diagnosis of FH. In the remainder, a polygenic aetiology is most likely, due to the co-inheritance of common LDL-C-raising variants. The cardiovascular presentation and management of FH will differ between patients based on their underlying genetic factors. Summary New genotyping methods such as next-generation sequencing will provide us with better understanding of the genetic architecture of FH.

Ana Catarina Alves, colaboradora do grupo de investigação cardiovascular do Departamento da Promoção da Saúde e Prevenção de Doenças Não Transmissíveis do INSA, IP e aluna de doutoramento da Faculdade de Ciências da Universidade de Lisboa recebeu com este estudo, no dia 2 de junho de 2014, o Prémio Jovem Investigador atribuído pela Sociedade Europeia de Aterosclerose. Familial hypercholesterolaemia (FH) is characterized by increased circulating low-density lipoprotein (LDL) cholesterol leading to premature atherosclerosis and coronary heart disease. Although FH is usually caused by mutations in LDLR, mutations in APOB and PCSK9 also cause FH but only a few mutations have been reported, APOB p.R3527Q being the most common. However, 30-80% of clinical FH patients do not present an identifiable mutation in any of the described genes. To identify the genetic cause of the hypercholesterolaemia in 65 patients without mutations in LDLR, PCSK9 or in fragments of exon 26 and 29 of APOB currently analysed, we performed whole sequencing of APOB by pyrosequencing. A total of 10 putative mutations in APOB were identified. Flow cytometry with fluorescently labelled LDL from patients and relatives showed that p.Arg1164Thr (exon 22) and p.Gln4494del (exon 29) presented a 40% decrease in internalization in lymphocytes and HepG2 cells, very similar to APOB3527. The proliferation assays with U937 cells showed reduced growth for both cases. The variant p.Tyr1247Cys was found to be neutral and other three alterations were considered polymorphisms. Our results emphasize the need to study the whole APOB in routine protocols to improve patient identification and cardiovascular risk assessment.

Background The most frequent monogenic causes of low plasma cholesterol are familial hypobetalipoproteinemia (FHBL1) because of truncating mutations in apolipoprotein B coding gene (APOB) and familial combined hypolipidemia (FHBL2) due to loss-of-function mutations in ANGPTL3 gene. Objective A direct comparison of lipid phenotypes of these 2 conditions has never been carried out. In addition, although an increased prevalence of liver steatosis in FHBL1 has been consistently reported, the hepatic consequences of FHBL2 are not well established. Methods We investigated 350 subjects, 67 heterozygous carriers of APOB mutations, 63 carriers of the p.S17* mutation in ANGPTL3 (57 heterozygotes and 6 homozygotes), and 220 noncarrier normolipemic controls. Prevalence and degree of hepatic steatosis were assessed by ultrasonography. Results A steady decrease of low-density lipoprotein cholesterol levels were observed from heterozygous to homozygous FHBL2 and to FHBL1 individuals, with the lowest levels in heterozygous FHBL1 carrying truncating mutations in exons 1 to 25 of APOB (P for trend

Role of theAPOBGene Polymorphism (c.12669G>A, p. Gln4154Lys) in Coronary Artery Disease in the Indian Punjabi PopulationHigh concentration of apolipoprotein B (apoB) is a risk factor for coronary artery disease (CAD). The association of theAPOBgene polymorphism c.12669G>A, p. Gln4154Lys with the risk of CAD varies considerably in different populations. The present study represents the first investigation regarding the role of thisAPOBgene polymorphism with CAD in the Indian Punjabi population. We have studied theAPOBgene polymorphism c.12669G>A, p. Gln4154Lys and its relationship with lipid, apoB, low-density lipoprotein (LDL) heterogeneity and oxidation in subjects suffering from CAD. The study was conducted on 87 patients with CAD; 75 healthy subjects served as controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to determine the DNA polymorphism in theAPOBgene. Frequency of R-(mutant) allele was significantly high (pAPOBgenotypes were insignificant (p>0.05). However, serum apoB levels were significantly raised (pAPOBgenotype. Coronary artery disease patients had raised significantly raised (pAPOBgenotype as compared to controls. Carriers of the R- allele are at higher risk of CAD, probably because of elevated serum apoB levels in the Indian Punjabi population. Overall, it may be concluded that the R- allele might be associated with increased susceptibility towards CAD development in the Indian Punjabi population, and one of the linking factor is the elevation in serum apoB levels. However, this association needs further evaluation in a larger population. Secondly, the robust mechanism behind the positive association of the R- allele with raised serum apoB levels needs to be explored, which might be helpful in the strengthening the observed results.

Background Familial hypercholesterolemia (FH) is a monogenic disorder of lipoprotein metabolism leading to an increased risk of premature cardiovascular disease. Genetic testing for FH is not commonly used in Asian countries. We aimed to define the genetic spectrum of FH in Hong Kong and to test the feasibility of cascade genetic screening. Methods Ninety‐six Chinese subjects with a clinical diagnosis of FH were recruited, and family‐based cascade screening incorporating genetic testing results was performed. Results Forty‐two distinct mutations were identified in 67% of the index FH cases. The majority of causative mutations were in the LDLR gene. The three commonest mutations in the LDLR gene were NM_000527.4(LDLR): c.1241 T>G, NM_000527.4(LDLR): c.1474G>A, and NM_000527.4(LDLR): c. 682G>A, and nine novel variants were identified. The NM_000384.2(APOB): c.10579 C>T variant of the APOB gene was found in 5% of the index subjects. The presence of causative mutation significantly increased the odds of successful family recruitment for screening with an OR of 3.7 (95% CI: 1.53–9.11, p = 0.004). Conclusion Approximately two‐third of the subjects in this clinically ascertained sample of patients with FH had a discrete genetic basis. Genetic identification improves the response rate and efficiency of family screening.

Background Homozygous familial hypercholesterolemia is a rare clinical phenotype with a variable expression, which is characterized by extremely elevated plasma low-density lipoprotein (LDL), tendon and skin xanthomas, and a progressive atherosclerosis. In 95% of patients, homozygous familial hypercholesterolemia is due to mutations in low-density lipoprotein receptor ( LDLR ) gene, which abolish (receptor-negative) or greatly reduce (receptor-defective) LDLR function. Objective The objective of the study was the molecular and phenotypic characterization of 4 siblings with severe hypercholesterolemia. Methods The major LDL-related genes ( LDLR , APOB , PCSK9 , ANGPTL3 , APOE , and APOC3 ) were sequenced. LDLR messenger RNA, isolated from leukocytes, was reverse transcribed and sequenced. Results The index cases were 24-year-old identical twin sisters with long-standing tendon xanthomas and high low-density lipoprotein cholesterol (LDL-C ∼10 mmol/L) but no coronary heart disease. They were carriers of 2 LDLR mutations: (1) a previously reported mutation [p.(G335S)] inherited from the mother who had LDL-C level within normal range; (2) a novel 24 bp deletion in exon 8/intron 8 junction inherited from the hypercholesterolemic (LDL-C 6.1 mmol/L) father. The deletion allele encodes an messenger RNA with a partial deletion of exon 8, whose translation product has an in-frame deletion of 17 amino acids [p.(Glu380_Gly396del)]. Family screening revealed that the 2 siblings of the twin sisters were also compound heterozygotes but had much lower LDL-C levels (8.2 and 7.1 mmol/L). The sequence of potential modifying genes showed that the 2 siblings and the mother of the twin sisters were heterozygous for a rare missense variant of apoB [p.(S2429T)], which might have an LDL-lowering effect. Conclusions We report a rare event of 4 siblings found to be compound heterozygotes for 2 LDLR gene mutations but showing a different phenotype severity. The less severely affected siblings were carriers of a rare apoB missense variant.

Monogenic hypercholesterolemia is a group of lipid disorders, most of which have autosomal dominant transmission. Familial defective apoB (FDB) resulting from mutations in the APOB gene is a well-recognized cause of autosomal dominant monogenic hypercholesterolemia (ADMH). However, the frequency of FDB among patients with ADMH is not well established. The aim of our research was to screen for mutations responsible for FDB in subjects with a clinical diagnosis of familial hypercholesterolemia. We studied 408 patients from the Spanish Register of Familial Hypercholesterolemia, proportionally distributed among all Spanish regions. Abnormal SSCP patterns of the APOB gene were checked by DNA sequencing and restriction analysis. Three out of the 408 patients were carriers of the R3500Q mutation, and 2 subjects were carriers of the silent T3552T mutation; in both of these patients functional mutations in the LDL receptor gene were found. We conclude that FDB is not a common cause of ADMH in Spain; the R3500Q mutation is the only mutation in APOB causing FDB, and the LDL receptor binding domain of APOB is highly conserved in the studied sample.

Familial hypobetalipoproteinemia (FHBL), an autosomal co-dominant disorder, is associated with reduced plasma concentrations (5th percentile for age and sex) of apolipoprotein (apo) B and beta-migrating lipoproteins. To date, only mutations in APOB encoding prematurely truncated apoB have been found in FHBL. We discovered a novel APOB gene mutation, namely R463W, in an extended Christian Lebanese FHBL kindred. Heterozygotes for R463W had the typical FHBL phenotype, whereas homozygotes had barely detectable apoB-100. The effect of the R463W mutation on apoB secretion was examined using transfected McA-RH7777 cells that expressed one of two recombinant human apoBs, namely B48 and B17. In both cases, the mutant proteins (B48RW and B17RW) were retained within the endoplasmic reticulum and were secreted poorly compared with their wild-type counterparts. Pulse-chase analysis showed that secretion efficiencies of B48RW and B17RW were, respectively, 45 and 40% lower than those of the wild-types. Substitution of Arg(463) with Ala in apoB-17 (B17RA) decreased secretion efficiency by approximately 50%, but substitution with Lys (B17RK) had no effect on secretion, indicating that the positive charge was important. Molecular modeling of apoB predicted that Arg(463) was in close proximity to Glu(756) and Asp(456). Substitution of Glu(756) with Gln (B17EQ) had no effect on secretion, but substitution of Asp(456) with Asn (B17DN) decreased secretion to the same extent as B17RW. In co-transfection experiments, the mutant B17RW showed increased binding to microsomal triglyceride transfer protein as compared with wild-type B17. Thus, the naturally occurring R463W mutant reveals a key local domain governing assembly and secretion of apoB-containing lipoproteins.

The present paper informs about allele frequency distributions for the two VNTR loci (ApoB, D1S80) in Slovak Romanies from Jarovnice and Richnava settlements. The results are compared with the data of non-Romany Slovak population. Thirteen segregating alleles were detected in a total sample of 267 Romany individuals on the locus ApoB. The most frequent alleles were 37 and 35 on the locus for ApoB gene. Genotype 37/37 occurs as the most frequent in Slovak Romany population. 13 alleles were detected in a total sample of 198 Romany individuals on the locus D1S80. Alleles 24 and 18 and genotype 24/24 were the most frequent in Romany individuals.

Familial hypobetalipoproteinemia (FHBL) is commonly caused by mutations in the apolipoprotein B gene ( APOB ). The APOB gene encodes 2 proteins, apolipoprotein (apo) B-48 and apo B-100. Apo B-48 is formed in the intestine and is essential for the formation and recognition of dietary derived chylomicrons, and apo B-100 is found in VLDLs and LDLs of hepatic origin and is involved in the endogenous transport of triglycerides, cholesterol, and fat-soluble vitamins. A number of abnormally truncated apo B proteins have been described, and by convention are referred to by a centile system reflecting their apparent M r in relation to apo B-100 (1). Truncations shorter than apo B-27 are not expressed in lipoproteins, and those shorter than apo B-75 are underrepresented in LDL (2)(3)(4). Consequently homozygous mutations in the N-terminal third of APOB result in the virtual absence of both apo B-48 and apo B-100 and their corresponding lipoproteins, and thus very low concentrations of plasma triglycerides, cholesterol, and the fat-soluble vitamins. This condition …

We have analyzed allele frequency distribution at the hypervariable locus 3 to the apolipoprotein B gene in a healthy population sample (241 women and 246 men) from the Belgrade area. The bimodal distribution of sixteen different hypervariable region (HVR) alleles and the heterozygosity index (average 0.76) in both samples are similar to ones reported for other Caucasian populations. However, we found the hypervariable element containing 34 repeats-HVE34 allele to be the most common one in both female and male samples, and that there was a lower frequency of the HVE GT 36 alleles. For further analysis of 3HVR allele frequency and its possible association with serum lipid levels, all HVR alleles were grouped into five main types (HVE LT 32, HVE32, HVE34, HVE36, and HVE GT 36), and the samples were divided into lipid quartiles. We found that (1) apo B gene 3HVR polymorphism is associated with variation of normal lipid levels in males, (2) HVE32 alleles are associated with decreased serum levels of total cholesterol, LDL cholesterol, and triglycerides, and (3) HVR genotypes containing HVE34 and HVE36 alleles are associated with elevated serum levels of total cholesterol, low density lipoprotein (LDL) cholesterol, and triglycerides. (C) 1998 Wiley-Liss, Inc.

Familial hypobetalipoproteinemia (FHBL) exists in three forms: a) FHBL genetically linked to truncated forms of apolipoprotein B (apoB); b) FHBL linked to the apoB gene but with no apoB truncations; and c) FHBL not linked to the apoB gene. Mean production rate (PR) of apoB-100 in FHBL subjects heterozygous for apoB truncations is approximately 30% of normal. In a 49-member D-kindred (FHBL phenotype defined as apoB < 40 mg/dl), no apoB truncations were detectable either by immunoblotting of plasma or by sequencing of relevant stretches of the apoB gene. Herein we report on the kinetic parameters of apoB-100-containing lipoproteins in four affected members of the D-kindred, and compare their kinetic values to 14 normal subjects, and 8 previously reported FHBL subjects heterozygous for various truncated forms of apoB. After an 8-h primed intravenous infusion of [13C]-leucine, enrichments of apoB-100 were assessed by gas chromatography-mass spectrometry and kinetic parameters were calculated by multicompartmental modeling. The affected members of the D-kindred had similar very low, intermediate, and low density lipoprotein (VLDL, IDL, and LDL) PRs as normal controls, but their fractional catabolic rates (FCR) for VLDL and LDL were approximately 2 and 3 times higher, respectively, than those of normals. By contrast in apoB truncation subjects, apoB-100 PRs were uniformly reduced, while apoB-100 FCRs were similar to normals. Thus, diverse physiologic mechanisms are responsible for the low apoB levels in these two different, genetically determined forms of FHBL.

Sudden death constitutes one of the most frequent causes of death in adults. In order to look for genetic markers that could play a role in the sudden death in adults (SDA), the common deletions of 4977 bp in the mitochondrial DNA and variations in the apoB gene were studied. The correlation of mtDNA common deletion with SDA victims might offer a rapid and simple tool to provide additional information in complex SDA investigations

Familial defective apolipoprotein B (FDB) is an autosomal dominant genetic disorder causing hypercholesterolemia in affected patients. It is occurring due to mutation of apoB gene leading to a decreased low density lipoprotein (LDL) particles clearance. The R3527Q mutation is one of the disease’s causative mutations. Data support mutation’s origin 6000-7000 years ago in Central Europe and its prevalence is decreasing in relation to the distance from where initially occurred. Until now, the presence of R3527Q mutation had not been confirmed in Greece. This is the first report of FDB due to R3527Q mutation in a family in Northwest Greece.

BACKGROUND: In the Netherlands, a screening programme was set up in 1994 in order to identify all patients with familial hypercholesterolaemia (FH). After 15 years of screening, we evaluated the geographical distribution, possible founder effects and clinical phenotype of the 12 most prevalent FH gene mutations. METHODS: Patients who carried one of the 12 most prevalent mutations, index cases and those identified between 1994 and 2009 through the screening programme and whose postal code was known were included in the study. Low-density lipoprotein cholesterol (LDL-C) levels at the time of screening were retrieved. The prevalence of identified FH patients in each postal code area was calculated and visualised in different maps. RESULTS: A total of 10,889 patients were included in the study. Mean untreated LDL-C levels ranged from 4.4 to 6.4 mmol/l. For almost all mutations, a region of high prevalence could be observed. In total, 51 homozygous patients were identified in the Netherlands, of which 13 true homozygous for one of the 12 most prevalent mutations. The majority of them were living in high-prevalence areas for that specific mutation. CONCLUSIONS: Phenotypes with regard to LDL-C levels varied between the 12 most prevalent FH mutations. For most of these mutations, a founder effect was observed. Our observations can have implications with regard to the efficiency of molecular screening and physician's perception of FH and to the understanding of the prevalence and distribution of homozygous patients in the Netherlands.

Familial hypobetalipoproteinemia (FHBL) is a co-dominant disorder characterized by reduced plasma levels of low density lipoprotein cholesterol (LDL-C) and its protein constituent apolipoprotein B (apoB), which may be due to mutations in APOB gene, mostly located in the coding region of this gene. We report two novel APOB gene mutations involving the acceptor splice site of intron 11 (c.1471-1G>A) and of intron 23 (c.3697-1G>C), respectively, which were identified in two patients with heterozygous FHBL associated with severe fatty liver disease. The effects of these mutations on APOB pre-mRNA splicing were assessed in COS-1 cells expressing the mutant APOB minigenes. The c.1471-1G>A APOB minigene generated two abnormal mRNAs. In one mRNA the entire intron 11 was retained; in the other mRNA exon 11 joined to exon 12, in which the first nucleotide was deleted due to the activation of a novel acceptor splice site. The predicted products of these mRNAs are truncated proteins of 546 and 474 amino acids, designated apoB-12.03 and apoB-10.45, respectively. The c.3697-1G>C APOB minigene generated a single abnormal mRNA in which exon 23 joined to exon 25, with the complete skipping of exon 24. This abnormal mRNA is predicted to encode a truncated protein of 1220 amino acids, designated apoB-26.89. These splice site mutations cause the formation of short truncated apoBs, which are not secreted into the plasma as lipoprotein constituents. This secretion defect is the major cause of severe fatty liver observed in carriers of these mutations.

Homozygous familial hypobetalipoproteinaemia (Ho-FHBL) is a rare co-dominant disorder characterized by extremely low levels of low-density lipoprotein cholesterol (LDL-C) and apolipoprotein B (apoB). Most patients with Ho-FHBL have mutations in APOB gene resulting in truncated apoBs. Some patients are asymptomatic, while others have fatty liver, intestinal fat malabsorption and neurological dysfunctions. We investigated three adult subjects with severe hypobetalipoproteinaemia and a family history of FHBL. Proband FHBL-47 had liver cirrhosis with hepatocarcinoma and a renal carcinoma but no clinical manifestations related to FHBL. He was a compound heterozygote for a 7-bp deletion in exon 21 and a base insertion in exon 26 resulting in truncated apoBs (apoB-22.46/apoB-66.51). Proband FHBL-53, with severe hepatic steatosis and fibrosis, had a nonsense mutation in exon 19 resulting in a truncated apoB (apoB-20.61) and a rare nucleotide substitution in intron 14 (c.2068-4T>A). The latter was also present in her daughter, found to have low plasma LDL-C and apoB. Proband FHBL-82 had chronic diarrhoea and steatorrhoea. She was found to be homozygous for a nonsense mutation in exon 24 resulting in a truncated apoB (apoB-26.65). In adult subjects, the presence of chronic liver disease and chronic diarrhoea, when associated with severe hypobetalipoproteinaemia, may lead to the diagnosis of Ho-FHBL.

Primary hypobetalipoproteinemia (HBL) includes a group of genetic disorders: abetalipoproteinemia (ABL) and chylomicron retention disease (CRD), with a recessive transmission, and familial hypobetalipoproteinemia (FHBL) with a co-dominant transmission. ABL and CRD are rare disorders due to mutations in the MTP and SARA2 genes, respectively. Heterozygous FHBL is much more frequent. FHBL subjects often have fatty liver and, less frequently, intestinal fat malabsorption. FHBL may be linked or not to the APOB gene. Most mutations in APOB gene cause the formation of truncated forms of apoB which may or may be not secreted into the plasma. Truncated apoBs with a size below that of apoB-30 are not detectable in plasma; they are more frequent in patients with the most severe phenotype. Only a single amino acid substitution (R463W) has been reported as the cause of FHBL. Approximately 50% of FHBL subjects are carriers of pathogenic mutations in APOB gene; therefore, a large proportion of FHBL subjects have no apoB gene mutations or are carriers of rare amino acid substitutions in apoB with unknown effect. In some kindred FHBL is linked to a locus on chromosome 3 (3p21) but the candidate gene is unknown. Recently a FHBL plasma lipid phenotype was observed in carriers of mutations of the PCSK9 gene causing loss of function of the encoded protein, a proprotein convertase which regulates LDL-receptor number in the liver. Inactivation of this enzyme is associated with an increased LDL uptake and hypobetalipoproteinemia. HBL carriers of PCSK9 mutations do not develop fatty liver disease.

Apolipoprotein B (apoB) gene expression is regulated by transcriptional, posttranscriptional, and translational mechanisms in both small intestinal enterocytes and hepatocytes of all mammals. This review focuses on the posttranscriptional regulation of apoB gene expression—specifically, an enzymatic modification to the nuclear transcript referred to as C to U RNA editing. C to U RNA editing, the process responsible for the production of apoB-48, is an exquisitely precise process that targets a single nucleotide in over 14,000. RNA editing requires regulated interactions between core enzyme components whose identity is known and regulatory factors, most of which have yet to be fully characterized. This review summarizes information relevant to those transacting factors that are essential for editing activity (core factors) as well as those interacting partners (regulatory factors) whose physiological role in RNA editing is yet to be fully defined.

Publisher Summary In 1986, Young et al. reported the presence of a truncated apolipoprotein (apo) B in the plasma of the members of kindred with hypobetalipoproteinemia. Subsequent studies revealed that the truncated apoB was caused by a frameshift mutation in the apoB gene. Since then, it has become apparent that a variety of apoB gene mutations that interfere with the translation of a full-length apoB-100 molecule can cause familial hypobetalipoproteinemia. Many of these mutations result in the production of a truncated species of apoB that can be detected within the plasma lipoproteins. A comprehensive review of familial hypobetalipoproteinemia that includes a summary of the historical aspects of the syndrome, clinical descriptions of the heterozygous and homozygous forms of the disorder, a list of apoB gene mutations, causing hypobetalipoproteinemia, and a summary of the properties of lipoproteins, containing truncated apoB species has been published. This chapter describes the techniques that are useful in identification and characterization of truncated apoB species.

INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 40 Transport of Exogenous (Dietary) Li pids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 40 Transport of Endogenous Lipids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 4 1 HDL and Reverse Cholesterol Transport . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 4 1 APOLIPOPROTEIN GENES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 42 Chromosomal Localization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 42 Evolutionary Considerations .. . . . . . . . . ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 42 APOLIPOPROTEIN GENE VARIATION . . . . . . . . .. . . .. . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . 1 45 Meth ods to Study Varia bility of Apolipoprotein Genes... . . . . . . . . . . . . 1 45 ApoA-/IC-III1A-/V Gene Cluster. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 46 ApoA-1I Gene . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 48 ApoB Gene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 49 ApoC-lI Gene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 52 ApoE Gene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 53 Apo(a ) Gene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155 ApoD Gene. . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155 CONCLUDING REMARKS . . . . . . . . . . . . . . . .... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 55