Leiden University Scholarly Publications

Zada, A.; Zhao, Y.Y.; Halim, D.; Windster, J.; Linde, H.C. van der; Glodener, J.; ... ; Alves, M.M. 2022

The long Filamin-A isoform is required for intestinal development and motility: implications for chronic intestinal pseudo-obstruction

Article / Letter to editor

open access

Filamin A (FLNA) is a cytoplasmic actin binding protein, recently shown to be expressed as a long and short isoform. Mutations in FLNA are associated with a wide spectrum of disorders, including an... Show moreFilamin A (FLNA) is a cytoplasmic actin binding protein, recently shown to be expressed as a long and short isoform. Mutations in FLNA are associated with a wide spectrum of disorders, including an X-linked form of chronic intestinal pseudo-obstruction (CIPO). However, the role of FLNA in intestinal development and function is largely unknown. In this study, we show that FLNA is expressed in the muscle layer of the small intestine from early human fetal stages. Expression of FLNA variants associated with CIPO, blocked expression of the long flna isoform and led to an overall reduction of RNA and protein levels. As a consequence, contractility of human intestinal smooth muscle cells was affected. Lastly, our transgenic zebrafish line showed that the flna long isoform is required for intestinal elongation and peristalsis. Histological analysis revealed structural and architectural changes in the intestinal smooth muscle of homozygous fish, likely triggered by the abnormal expression of intestinal smooth muscle markers. No defect in the localization or numbers of enteric neurons was observed. Taken together, our study demonstrates that the long FLNA isoform contributes to intestinal development and function. Since loss of the long FLNA isoform does not seem to affect the enteric nervous system, it likely results in a myopathic form of CIPO, bringing new insights to disease pathogenesis. Show less

Houlleberghs, H.; Dekker, M.; Lusseveld, J.; Pieters, W.; Ravesteyn, T. van; Verhoef, S.; ... ; Riele, H. te 2020

Three-step site-directed mutagenesis screen identifies pathogenic MLH1 variants associated with Lynch syndrome

Article / Letter to editor

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BackgroundInactivating mutations in the MLH1 DNA mismatch repair (MMR) gene underlie 42% of Lynch syndrome (LS) cases. LS is a cancer predisposition causing early onset colorectal and endometrial... Show moreBackgroundInactivating mutations in the MLH1 DNA mismatch repair (MMR) gene underlie 42% of Lynch syndrome (LS) cases. LS is a cancer predisposition causing early onset colorectal and endometrial cancer. Nonsense and frameshift alterations unambiguously cause LS. The phenotype of missense mutations that only alter a single amino acid is often unclear. These variants of uncertain significance (VUS) hinder LS diagnosis and family screening and therefore functional tests are urgently needed. We developed a functional test for MLH1 VUS termed 'oligonucleotide-directed mutation screening' (ODMS).MethodsThe MLH1 variant was introduced by oligonucleotide-directed gene modification in mouse embryonic stem cells that were subsequently exposed to the guanine analogue 6-thioguanine to determine whether the variant abrogated MMR.ResutsIn a proof-of-principle analysis, we demonstrate that ODMS can distinguish pathogenic and non-pathogenic MLH1 variants with a sensitivity of >95% and a specificity of >91%. We subsequently applied the screen to 51 MLH1 VUS and identified 31 pathogenic variants.ConclusionODMS is a reliable tool to identify pathogenic MLH1 variants. Implementation in clinical diagnostics will improve clinical care of patients with suspected LS and their relatives. Show less

Goverde, A.; Spaander, M.C.W.; Nieboer, D.; Ouweland, A.M.W. van den; Dinjens, W.N.M.; Dubbink, H.J.; ... ; Wagner, A. 2018

Evaluation of current prediction models for Lynch syndrome: updating the PREMM5 model to identify PMS2 mutation carriers

Article / Letter to editor

open access

Kinnersley, B.; Chubb, D.; Dobbins, S.E.; Frampton, M.; Buch, S.; Timofeeva, M.N.; ... ; Houlston, R.S. 2016

Correspondence: SEMA4A variation and risk of colorectal cancer

Article / Letter to editor

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Rasmussen, L.J.; Heinen, C.D.; Royer-Pokora, B.; Drost, M.; Tavtigian, S.; Hofstra, R.M.W.; Wind, N. de 2012

Pathological assessment of mismatch repair gene variants in Lynch syndrome: Past, present, and future

Article / Letter to editor

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Drost, M.; Zonneveld, J.B.; Hees, S. van; Rasmussen, L.J.; Hofstra, R.M.W.; Wind, N. de 2011

A rapid and cell-free assay to test the activity of lynch syndrome-associated MSH2 and MSH6 missense variants

Article / Letter to editor

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Lynch syndrome (LS) is an autosomal dominant disorder that predisposes to colon, endometrial, and other cancers. LS is caused by a heterozygous germline mutation in one of the DNA mismatch repair ... Show moreLynch syndrome (LS) is an autosomal dominant disorder that predisposes to colon, endometrial, and other cancers. LS is caused by a heterozygous germline mutation in one of the DNA mismatch repair (MMR) genes. A significant proportion of all mutations found in suspected LS patients comprises single amino acid alterations. The pathogenicity of these variants of uncertain significance (VUS) is difficult to assess, precluding diagnosis of carriers and their relatives. Here we present a rapid cell-free assay to investigate MMR activity of MSH2 or MSH6 VUS. We used this assay to analyze a series of MSH2 and MSH6 VUS, selected from the Leiden Open Variation Database. Whereas a significant fraction of the MSH2 VUS has lost MMR activity, suggesting pathogenicity, the large majority of the MSH6 VUS appears MMR proficient. We anticipate that this assay will be an important tool in the development of a comprehensive and widely applicable diagnostic procedure for LS-associated VUS. Hum Mutat 00:1-7, 2011. © 2011 Wiley Periodicals, Inc. Show less

Vos, Y.J.; Walle, H.E.K. de; Bos, K.K.; Stegeman, J.A.; Berge, A.M. ten; Bruining, M.; ... ; Hofstra, R.M.W. 2010

Genotype-phenotype correlations in L1 syndrome: a guide for genetic counselling and mutation analysis

Article / Letter to editor

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Objectives To develop a comprehensive mutation analysis system with a high rate of detection, to develop a tool to predict the chance of detecting a mutation in the L1CAM gene, and to look for... Show moreObjectives To develop a comprehensive mutation analysis system with a high rate of detection, to develop a tool to predict the chance of detecting a mutation in the L1CAM gene, and to look for genotype-phenotype correlations in the X-linked recessive disorder, L1 syndrome. Methods DNA from 367 referred patients was analysed for mutations in the coding sequences of the gene. A subgroup of 100 patients was also investigated for mutations in regulatory sequences and for large duplications. Clinical data for 106 patients were collected and used for statistical analysis. Results 68 different mutations were detected in 73 patients. In patients with three or more clinical characteristics of L1 syndrome, the mutation detection rate was 66% compared with 16% in patients with fewer characteristics. The detection rate was 51% in families with more than one affected relative, and 18% in families with one affected male. A combination of these two factors resulted in an 85% detection rate (OR 10.4, 95% CI 3.6 to 30.1). The type of mutation affects the severity of L1 syndrome. Children with a truncating mutation were more likely to die before the age of 3 than those with a missense mutation (52% vs 8%; p=0.02). Conclusions We developed a comprehensive mutation detection system with a detection rate of almost 20% in unselected patients and up to 85% in a selected group. Using the patients' clinical characteristics and family history, clinicians can accurately predict the chance of finding a mutation. A genotype-phenotype correlation was confirmed. The occurrence of (maternal) germline mosaicism was proven. Show less