Deletion mutation in BSCL2 gene underlies congenital generalized lipodystrophy in a Pakistani family

Congenital generalized lipodystrophy (CGL), or Berardinelli-Seip congenital lipodystrophy (BSCL), is a genetically heterogeneous disorder characterized by loss of adipose tissues and sub-cutaneous fats, enlarged fatty liver, hypertrophic muscles, Acanthosis nigricans, increased serum triglyceride level, insulin intolerance or diabetes mellitus. Pathogenicity of four genes has been reported in four clinically overlapping phenotypes so far.

The affected individuals of congenital generalized lipodystrophy type 1 (CGL1, MIM 608594) are characterized by typical poor fat accumulation in the metabolically active and mechanical adipose tissues. In addition these patients have special features of lytic bone lesions that are absent in other forms of CGL [1, 2]. Loss-of-function mutations in 1-acylglycerol-3-phosphate-O-Acyltransferase (AGPAT2, MIM 603100) gene at chromosome 9q34.3 have been assigned to cause CGL1 phenotype [3, 4].

Pathogenic variants of homologous to mouse gamma-3-linked (BSCL2, MIM 606158) gene, located at chromosome 11q13, cause congenital generalized lipodystrophy type 2 (CGL2, MIM 269700) or a seipin-deficient phenotype [3, 5]. The subcutaneous biopsies examinations of patients have revealed scattered groups of small adipocytes with low but detectable lipid content [6]. The seipin-deficient individuals have generalized congenital lipodystrophy, hypertrophic cardiomyopathy, higher rates of mild mental retardation and an earlier onset of diabetes [7–9].

Mutations in caveolin 1 (CAV1, MIM 601047) gene, located at chromosome 7q31.3, cause congenital generalized lipodystrophy type 3 (CGL3, MIM 612526). Caveolin-1-deficient patients exhibit unique manifestation of short stature, hypocalcemia and vitamin D resistance [10]. Recently another form congenital generalized lipodystrophy type 4 (CGL4, MIM 613327) has also been described caused by mutations in RNA polymerase 1 and transcript release factor (PTRF, MIM 603198) gene at chromosome 17q21 [11].

In the present study clinical and molecular analysis of a four generations consanguineous Pakistani family demonstrating autosomal recessive CGL phenotype was performed. Direct sequencing of four candidate genes in a Pakistani family revealed a single base pair deletion mutation in BSCL2 gene.

Loss-of-function mutations in the BSCL2 gene cause a severe form of lipodystrophy, whilst characteristic gain-of-function mutations are believed to be associated with aggregation of unfolded protein in endoplasmic reticulum resulting in neurodegeneration leading to a heterogeneous group of neuropathies [5]. The study presented here was a four generations consanguineous family, originated from Peshawar city Pakistan. Clinical features of the affected individuals resembled CGL2 phenotype. Moreover the first degree consanguinity of the parents and autosomal recessive mode of inheritance of the disease phenotype was a definite clue for a homozygous mutation. Mutations in BSCL2 leading to CGL2 phenotype have been identified worldwide [5, 14–16].

The generalized loss of adipose tissue, increased triglyceride levels and steatosis of the liver were comparable to a homozygous mutation p.Tyr213ThrfsX20 identified in an Indian family [5]. Early onset diabetes mellitus present in our patients were comparable with homozygous nonsense mutations identified in Chinese and severe insulin resistance in Japanese and Brazilian patients [9, 14, 15], however we did not perform the insulin resistance test in our patients. The hypertrophic cardiomyopathy identified in our cases was less severe as compared to Chinese patient caused by homozygous nonsense mutation [17]. There was mild mental retardation (IQ score 65–75) in both of our patients but the brain MRI did not reveal brain atrophy.

The variable features of neuropathies are mostly associated with heterozygous mutations in BSCL2 dHMN, Charcot-Marie-Tooth (CMT) and Silver syndromes [16, 18, 19]. In 2005, based on disease allele penetrance and severity of phenotype caused by heterozygous mutations in BSCL2 gene p.N88S in Austrian and German families, these patients were classified into six subclinical groups with overlapping features of HMN, CMT and Silver syndrome [20]. However, in 2009, Brusse et al. discovered a digenic inheritance of hereditary motor neuropathy in a large Dutch family with heterozygous mutation p.N88S in the BSCL2 gene and segregating autosomal dominant disease haplotype at chromosome 16p [21]. The phenotypic variability ranged from strictly neuropathic weakness to a spastic paraplegia with hereditary motor neuropathy presenting clinical phenotype similar to Silver syndrome [21]. Very recently another heterozygous mutation p.S90W identified in two Korean CMT type 2 disease patients [22] was associated with increased density of myelinated fibers. However, this feature was slightly different from the previously reported mutation p.S90L in three Italian patients representing CMT type 2 phenotype with pyramidal signs and subclinical sensory involvement on sural nerve biopsy [23].

Human BSCL2 gene encodes 398 or 462 amino acids seipin protein from either of the three transcripts 1.6 kb, 1.8 kb and 2.2 kb [24]. The 1.8 kb transcript is exclusively expressed in brain and testis while the other two are ubiquitously expressed [18]. Seipin protein is located in endoplasmic reticulum acting as a regulator of lipid catabolism and is essential for the differentiation of fat cells [18, 24]. Studies have revealed role of seipin in proper lipid storage and regulation of cAMP/PKA-mediated lipolysis in adipose differentiation [25, 26]. In adult mice brain bscl2 expression studies have suggested the possible involvement of seipin in central regulation of energy balance.

More than three hundred cases of BSCL have been reported in the medical literature with an estimated prevalence of 1 in 10 million people in USA [35]. However this condition is more common in other populations around the world like Lebanon, Brazil, Portugal and Sultanate of Oman with an estimated prevalence of 1:25000 to 1:1000000 [36]. The incidence data of BSCL and many other rare disorders are not available from Pakistan partly due to lack of disease registry database systems and non-availability of neonatal screening programs in the hospitals.

Generally, in families segregating autosomal recessive disorders, high frequency of consanguineous marriages may increase the frequency of homozygotes in the population leading to increased incidence of certain lesions, their founder effect and also the appearance of new mutations [37]. Although most causative mutations in Mendelian diseases are reported in single families, certain mutations may occur more frequently in some populations than others. Few examples of such prevalent mutations may be observed in Wilson’s disease [38], hemophilia [39], Hurler disease [37] and thalassemia [40]. Knowledge of the differences in the worldwide distribution of particular mutations may help to design shortcuts for genetic diagnosis and screening of relevant inherited diseases.

The identified mutation c.636delC (p.Tyr213ThrfsX20) in exon 5 of BSCL2 gene is the first report of CGL2 from Pakistan. The current study extends the body of evidence that describes the role of BSCL2 in congenital generalized lipodystrophy and association of loss-of-function mutations with severe lipodystrophy phenotype. The clinical features in our patients are similar to CGL2 phenotype but lack of motor neuropathies that are usually associated with Silver syndrome and Charcot-Marie-Tooth disease were not observed in our case. Understanding the molecular defects of BSCL2 gene in the patients can be helpful in the genetic counseling and prenatal diagnosis of affected families and may help to improve specific therapeutic interventions.