Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or . mil. Before sharing sensitive information, make sure you’re on a federal government site. VSports app下载.

Https

The site is secure. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. V体育官网.

. 2015 Mar 21:15:36.
doi: 10.1186/s12876-015-0261-y.

Congenital lactose intolerance is triggered by severe mutations on both alleles of the lactase gene

Lena Diekmann  1 , Katrin Pfeiffer  2 , Hassan Y Naim  3
Affiliations

V体育安卓版 - Congenital lactose intolerance is triggered by severe mutations on both alleles of the lactase gene

"VSports注册入口" Lena Diekmann et al. BMC Gastroenterol. .

Abstract

Background: Congenital lactase deficiency (CLD) is a rare severe autosomal recessive disorder, with symptoms like watery diarrhea, meteorism and malnutrition, which start a few days after birth by the onset of nursing. The most common rationales identified for this disorder are missense mutations or premature stop codons in the coding region of the lactase-phlorizin hydrolase (LPH) gene. Recently, two heterozygous mutations, c. 4419C > G (p. Y1473X) in exon 10 and c. 5387delA (p. D1796fs) in exon 16, have been identified within the coding region of LPH in a Japanese infant with CLD VSports手机版. .

Methods: Here, we investigate the influence of these mutations on the structure, biosynthesis and function of LPH V体育安卓版. Therefore the mutant genes were transiently expressed in COS-1 cells. .

Results: We show that both mutant proteins are mannose-rich glycosylated proteins that are not capable of exiting the endoplasmic reticulum. These mutant proteins are misfolded and turnover studies show that they are ultimately degraded. The enzymatic activities of these mutant forms are not detectable, despite the presence of lactase and phlorizin active sites in the polypeptide backbone of LPH-D1796fs and LPH-Y1473X respectively V体育ios版. Interestingly, wild type LPH retains its complete enzymatic activity and intracellular transport competence in the presence of the pathogenic mutants suggesting that heterozygote carriers presumably do not show symptoms related to CLD. .

Conclusions: Our study strongly suggests that the onset of severe forms of CLD is elicited by mutations in the LPH gene that occur in either a compound heterozygous or homozygous pattern of inheritance. VSports最新版本.

PubMed Disclaimer

Figures

Figure 1
Figure 1
Biosynthesis and transport kinetics of wild type and mutant proteins. COS-1 cells were transiently transfected with cDNAs encoding the wild type LPH, LPH-Y1473X or LPH-D1796fs and used 48 h after transfection. A) The cells were metabolically labeled with [35S] methionine for 6 h continuously, lysed, immunoprecipitated with mAb anti-LPH and treated with endo H. B) The cells were labeled with [35S] methionine for 2 h and chased for 0, 4, 8 and 12 h, followed by cell lysis and immunoprecipitation. In all these experiments, the proteins were subjected to SDS-PAGE and autoradiography. The quantification was performed by Quantity One® software.
Figure 2
Figure 2
Intracellular localization of wild type and the mutants in COS-1 cells. For visualization of LPH, transiently transfected COS-1 cells, expressing the wild type LPH, LPH-Y1473X or LPH-D1796fs, were fixed with 4% paraformaldehyde and permeabilised with 0.5% saponin. Immunolabeling was carried out using mAb anti-LPH (1:1000) as the primary antibody and anti-mouse IgG conjugated with Alexa 488 as the secondary antibody (1:500). The samples were analyzed by confocal laser microscopy.
Figure 3
Figure 3
Potential interaction of LPH-Y1473X and LPH-D1796fs with wild type LPH in co-transfection experiments. COS-1 cells were transiently transfected either with cDNA encoding wild type LPH tagged with myc (LPH-myc) or cDNA clones encoding the mutants LPH-Y1473X and LPH-D1796fs. In another set of experiments, coexpression of LPH-myc with LPH-Y1473X or with LPH-D1796fs was performed in COS-1 cells. 48 h posttransfection the cells were metabolically labeled with [35S] methionine for 6 h continuously, lysed and the lysates were immunoprecipitated with either an antibody against myc to detect wild type LPH or a mixture of mAb anti-LPH antibodies to detect the mutants. Lactase activity was measured by determining the concentration of glucose generated by lactose hydrolysis by the Glucose oxidase-peroxidase mono-reagent method using photometric analysis. The total protein amount of each sample was determined by SDS-PAGE and autoradiography to calculate subsequently the relative specific activity. The quantification was performed by Quantity One® software. The results of LPH-Y1473X were taken from another gel with higher quality.
Figure 4
Figure 4
Biosynthesis and glycosylation pattern of wild type LPH, LPH-Y1473X and LPH-D1796fs at 20°C and 37°C. Transiently transfected COS-1 cells, expressing wild type LPH, LPH-Y1473X or LPH-D1796fs were labeled with [35S] methionine for 3 h and chased for 6 h and 18 h at 20°C (A) and 37°C (B) followed by cell lysis, immunoprecipitation and endo H treatment. The proteins were subjected to SDS-PAGE and autoradiography. The quantification was performed by Quantity One® software.
See this image and copyright information in PMC

References

    1. Lifshitz F. Congenital lactase deficiency. J Pediatr. 1966;69:229–37. doi: 10.1016/S0022-3476(66)80325-6. - DOI (VSports在线直播) - PubMed
    1. Savilahti E, Launiala K, Kuitunen P. Congenital lactase deficiency. A clinical study on 16 patients. Arch Dis Child. 1983;58:246–52. doi: 10.1136/adc.58.4.246. - DOI - PMC - PubMed
    1. Uchida N, Sakamoto O, Irie M, Abukawa D, Takeyama J, Kure S, et al. Two novel mutations in the lactase gene in a Japanese infant with congenital lactase deficiency. Tohoku J Exp Med. 2012;227:69–72. doi: 10.1620/tjem.227.69. - V体育平台登录 - DOI - PubMed
    1. Torniainen S, Freddara R, Routi T, Gijsbers C, Catassi C, Hoglund P, et al. Four novel mutations in the lactase gene (LCT) underlying congenital lactase deficiency (CLD) BMC Gastroenterol. 2009;9:8. doi: 10.1186/1471-230X-9-8. - DOI - PMC - PubMed
    1. Behrendt M, Keiser M, Hoch M, Naim HY. Impaired trafficking and subcellular localization of a mutant lactase associated with congenital lactase deficiency. Gastroenterology. 2009;136:2295–303. doi: 10.1053/j.gastro.2009.01.041. - DOI (VSports在线直播) - PubMed

MeSH terms

V体育安卓版 - Supplementary concepts

VSports手机版 - LinkOut - more resources