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S Kenwrick

Publications and source records attributed to S Kenwrick.

At least 37 records · Page 2Linked to original sources
PubMed1998

Chinese hamster ovary cells resistant to the topoisomerase II catalytic inhibitor ICRF-159: a Tyr49Phe mutation confers high-level resistance to bisdioxopiperazines.

Anticancer drugs targeted to the nuclear enzyme DNA topoisomerase II are classified as poisons that lead to DNA breaks or catalytic inhibitors that appear to completely block enzyme activity. To examine the effects of the bisdioxopiperazine class of catalytic inhibitors to topoisomerase II, we investigated a Chinese hamster ovary (CHO) subline selected for resistance to ICRF-159 (CHO/159-1). Topoisomerase IIalpha content in CHO/159-1 cells was reduced by 40-50%, compared to wild-type CHO cells, whereas the beta isoform was increased by 10-20% in CHO/159-1 cells. However, the catalytic activity of topoisomerase II in nuclear extracts from CHO/159-1 cells was unchanged, as was its inhibition by the topoisomerase II poison etoposide (VP-16). No inhibition of topoisomerase II catalytic activity by ICRF-187 was seen in CHO/159-1 cells up to 500 microM, whereas inhibition was evident at 50 microM in wild-type CHO cells. VP-16-mediated DNA single-strand breaks and cytotoxicity were similar in the two sublines. ICRF-187 could abrogate these VP-16 effects in the wild-type line but had no effect in CHO/159-1 cells. Western blots of topoisomerase IIalpha after incubation of CHO cells with ICRF-187 demonstrated a marked band depletion, whereas this effect was completely lacking in CHO/159-1 cells, and an equal effect of VP-16 was observed in both lines. These data imply that the CHO/159-1 topoisomerase IIalpha lacks sensitivity to bisdioxopiperazines and that the mechanism of resistance in this cell line does not confer cross-resistance to topoisomerase II poisons, suggesting that mutations conferring resistance to bisdioxopiperazines can occur at sites distinct from those responsible for resistance to complex stabilizing agents. Accordingly, CHO/159-1 cDNA showed two heterozygous mutations in the proximal NH2-terminal part of topoisomerase IIalpha (Tyr49Phe and delta 309Gln-Gln-Ile-Ser-Phe313), which is in contrast to those induced by topoisomerase II poisons, which cluster further downstream. Site-directed mutagenesis and transformation of the homologous Tyr50Phe coding mutation in human topoisomerase IIalpha in a temperature-conditional yeast system demonstrated a high-level resistance to ICRF-193, compared to cells expressing wild-type cDNA, but none toward the poisons VP-16 or amsacrine, thus confirming that the Tyr50Phe mutation confers specific resistance to bisdioxopiperazines. Thus, these results indicate that the region of the protein involved in ATP-binding also plays a critical role in sensitivity to bisdioxopiperazines, a result consistent with the known requirement for the formation of an ATP-bound closed clamp for bisdioxopiperazine activity. These results may enable a more precise understanding of the interaction of topoisomerase II-directed drugs with their target enzyme.

Amino Acid Sequence↗
PubMed1998

The neural cell adhesion molecule L1: genomic organisation and differential splicing is conserved between man and the pufferfish Fugu.

The human gene for the neural cell adhesion molecule L1 is located on Xq28 between the ALD and MeCP2 loci. Mutations in the L1 gene are associated with four related neurological disorders, X-linked hydrocephalus, spastic paraplegia (SPG1), MASA syndrome, and X-linked corpus callosum agenesis. The clinical relevance of L1 has led us to sequence the L1 gene in human and to investigate its conservation in the vertebrate model genome of the pufferfish, Fugu rubripes (Fugu), a species with a compact genome of around 40Mb. For this purpose we have sequenced a human and a Fugu cosmid clone containing the corresponding L1 genes. For comparison, we have also amplified and sequenced the complete Fugu L1 cDNA. We find that the genomic structure of L1 is conserved. The human and Fugu L1 gene both have 28 exons of nearly identical size. Differential splicing of exons 2 and 27 is conserved over 430 million years, the evolutionary time span between the teleost Fugu and the human L1 gene. In contrast to previously published Fugu genes, many introns are larger in the Fugu L1 gene, making it slightly larger in size despite the compact nature of the Fugu genome. Homology at the amino acid and the nucleotide level with 40% and 51%, respectively, is lower than that of any previously reported Fugu gene. At the level of protein structure, both human and Fugu L1 molecules are composed of six immunoglobulin (Ig)-like domains and five fibronectin (Fn) type III domains, followed by a transmembrane domain and a short cytoplasmic domain. Only the transmembrane and the cytoplasmic domains are significantly conserved in Fugu, supporting their proposed function in intracellular signalling and interaction with cytoskeletal elements in the process of neurite outgrowth and fascicle formation. Our results show that the cytoplasmic domain can be further subdivided into a conserved and a variable region, which may correspond to different functions. Most pathological missense mutations in human L1 affect conserved residues. Fifteen out of 22 reported missense mutations alter amino acids that are identical in both species.

Alternative Splicing↗
PubMed1998

Somatic and germ line mosaicism and mutation origin for a mutation in the L1 gene in a family with X-linked hydrocephalus.

X-linked hydrocephalus is caused by mutations in the gene for neural cell adhesion molecule L1 (L1CAM). In this report, we describe identification of a mutation in an isolated case of hydrocephalus with adducted thumbs. Tracing the origin of the mutation within the family showed a degree of somatic mosaicism in the asymptomatic maternal grandfather of the propositus. This report highlights the need to take mosaicism into account when counselling relatives of affected individuals.

Cell Adhesion Molecules, Neuronal↗
PubMed1998

Neural cell adhesion molecule L1: relating disease to function.

Neural cell adhesion molecules of the immunoglobulin superfamily are important components of the network of guidance cues and receptors that govern axon growth and guidance during development. For neural cell adhesion molecule L1, the combined application of human genetics, knockout mouse technology, and cell biology is providing fundamental insight into the role of L1 in mediating neuronal differentiation. Disease-causing mutations as well as mouse models of L1 disruption can now be used to examine the relevance of L1 binding specificities and signal transduction pathways that have been observed in vitro.

Animals↗
PubMed1998

Alternative splicing of human NrCAM in neural and nonneural tissues.

Neural cell adhesion molecule NrCAM exists in a variety of isoforms as a result of alternative splicing of individual exons during RNA processing. In this report we demonstrate that many of the alternative splicing events described for chick are conserved in man and describe a novel variant of NrCAM cDNA. Furthermore, we show that NrCAM is expressed at significant levels outside the nervous system; in particular in pancreas, adrenal glands, and placenta and that expression in both brain and other tissues is accompanied by a very variable pattern of exon utilization in fetal and adult cells.

Adrenal Glands↗
PubMed1998

Neural cell recognition molecule L1: from cell biology to human hereditary brain malformations.

The neural cell recognition molecule L1 is a member of the immunoglobulin superfamily implicated in embryonic brain development. L1 is engaged in complex extracellular interactions, with multiple binding partners on cell surfaces and in the extracellular matrix. It is the founder of a neural group of related cell surface receptors that share with L1 a highly conserved cytoplasmic domain that associates with the cytoskeleton. Phenotypic analyses of human patients with mutations in the L1 gene and characterizations of L1-deficient mice suggest that L1 is important for embryonic brain histogenesis, in particular the development of axon tracts.

Animals↗
PubMed1997

Nine novel L1 CAM mutations in families with X-linked hydrocephalus.

Mutations in the gene for neural cell adhesion molecule L1 are responsible for the highly variable phenotype found in families with X-linked hydrocephalus, MASA syndrome, and spastic paraplegia type I. To date, 32 different mutations have been observed, the majority being unique to individual families. Here, we report nine novel mutations in L1 in 10 X-linked hydrocephalus families. Four mutations truncate the L1 protein and eliminate cell surface expression, and two would produce abnormal L1 through alteration of RNA processing. A further two of these mutations are small in-frame deletions that have occurred through a mechanism involving tandem repeated sequences. Together with a single missense mutation, these latter examples contribute to the growing number of existing mutations that affect short regions of the L1 protein that may have particular functional significance.

Amino Acid Sequence↗
PubMed1996

Outline structure of the human L1 cell adhesion molecule and the sites where mutations cause neurological disorders.

The L1 cell adhesion molecule has six domains homologous to members of the immunoglobulin superfamily and five homologous to fibronectin type III domains. We determined the outline structure of the L1 domains by showing that they have, at the key sites that determine conformation, residues similar to those in proteins of known structure. The outline structure describes the relative positions of residues, the major secondary structures and residue solvent accessibility. We use the outline structure to investigate the likely effects of 22 mutations that cause neurological diseases. The mutations are not randomly distributed but cluster in a few regions of the structure. They can be divided into those that act mainly by changing conformation or denaturing their domain and those that alter its surface properties.

Amino Acid Sequence↗
PubMed1996

X linked hydrocephalus and MASA syndrome.

X linked hydrocephalus and MASA syndrome are clinically related, neurological disorders with an X linked recessive mode of inheritance. Although originally described as distinct entities, their similarity has become apparent as the number of reported families has increased and a high degree of intra- and interfamilial variation in clinical signs noted for both disorders. Consideration of this clinical overlap together with finding that genes for both diseases map to the same chromosomal band (Xq28) led to the hypothesis that they were caused by mutation at the same locus. This was confirmed by identification of mutations in patients with X linked hydrocephalus and MASA syndrome within the gene for neural cell adhesion molecule L1. Here we review the clinical and genetic characteristics of these disorders and the underlying molecular defects in the L1 gene.

Abnormalities, Multiple↗
PubMed1996

Discordant segregation of Xq28 markers and a mutation in the L1 gene in a family with X linked hydrocephalus.

X linked recessive hydrocephalus is the most common hereditary form of hydrocephalus. Genetic analysis indicates that the majority of cases are caused by mutations in a single gene in Xq28, recently identified as the gene for neural cell adhesion molecule L1. Genetic heterogeneity for this disorder was suggested following the description of a single large pedigree where X linked hydrocephalus showed lack of linkage to Xq28 markers flanking the L1 gene. Mutation analysis in this family shows a single base pair deletion within the coding sequence of the L1 gene that would result in truncation of the mature protein. The nature of the mutation and its segregation with the disease through the pedigree indicate that it is the cause of X linked hydrocephalus in this family. These results are at odds with data obtained through segregation of alleles for markers flanking the L1 gene. Somatic and germline mosaicism is the most plausible explanation for these data, which also provide further evidence for genetic homogeneity of X linked hydrocephalus.

Base Sequence↗
PubMed1995

Gene analysis of L1 neural cell adhesion molecule in prenatal diagnosis of hydrocephalus.

X-linked hydrocephalus is the most common form of inherited hydrocephalus, and is associated with severe neurological deficits and premature death. We have shown that mutations in the gene encoding L1 neural cell adhesion molecule result in X-linked hydrocephalus, which enables improved prenatal diagnosis and investigation of the role of this molecule in sporadic cases. Here we report two pedigrees with apparently sporadic hydrocephalus in which we demonstrated a disabling mutation in the L1 gene. This enabled us to provide definitive prenatal diagnosis at 10 weeks' gestation.

Cell Adhesion Molecules, Neuronal↗
PubMed1995

Mutations in the cell adhesion molecule L1 cause mental retardation.

Recently, studies in the usually disparate fields of human genetics and developmental neurobiology have converged to reveal that some types of human mental retardation and brain malformations are due to mutations that affect the neural cell adhesion molecule L1. L1 has a very complex biology, interacting with a variety of ligands, and functioning in migration of neurons and growth of axons. Over the past few years, it has also become clear that L1 is able to influence intracellular second messengers. The identification of a number of different mutations in L1, some of which alter the extracellular portion of the molecule, and others that change only the cytoplasmic tail, confirm that L1 is a crucial player in normal brain development. The information gained from genetic analysis of human L1 is giving new insights into how L1 functions in the formation of major axon pathways, but it also raises unanticipated questions about how L1 participates in the development of cortical and ventricular systems.

Cell Adhesion Molecules↗
PubMed1995

Exon 2 of the gene for neural cell adhesion molecule L1 is alternatively spliced in B cells.

L1CAM is a neural cell adhesion molecule expressed mainly on neurones' cell surface and plays an important role in the developing fetal brain. Recently, we have shown that mutations in the gene encoding L1CAM are responsible for three related neurological disorders including the most common form of inherited hydrocephalus. During our genetic analysis, we have discovered that L1CAM is also expressed on the surface of B cells but that the messenger RNA in this tissue is different to that in brain through alternative splicing of the L1 gene. This indicates that this region of the L1 molecule has a distinct role in brain cells compared to B lymphocytes and confirms its importance in brain development.

Alternative Splicing↗
PubMed1995

New domains of neural cell-adhesion molecule L1 implicated in X-linked hydrocephalus and MASA syndrome.

The neural cell-adhesion molecule L1 is involved in intercellular recognition and neuronal migration in the CNS. Recently, we have shown that mutations in the gene encoding L1 are responsible for three related disorders; X-linked hydrocephalus, MASA (mental retardation, aphasia, shuffling gait, and adducted thumbs) syndrome, and spastic paraplegia type I (SPG1). These three disorders represent a clinical spectrum that varies not only between families but sometimes also within families. To date, 14 independent L1 mutations have been reported and shown to be disease causing. Here we report nine novel L1 mutations in X-linked hydrocephalus and MASA-syndrome families, including the first examples of mutations affecting the fibronectin type III domains of the molecule. They are discussed in relation both to phenotypes and to the insights that they provide into L1 function.

Abnormalities, Multiple↗
PubMed1994

A DNA fragment from Xq21 replaces a deleted region containing the entire FVIII gene in a severe hemophilia A patient.

In this paper we report the molecular characterization of a large deletion that removes the entire Factor VIII gene in a severe hemophilia A patient. Accurate DNA analysis of the breakpoint region revealed that a large DNA fragment replaced the 300-kb one, which was removed by the deletion. Pulsed-field gel electrophoresis analysis revealed that the size of the inserted fragment is about 550 kb. In situ hybridization demonstrated that part of the inserted region normally maps to Xq21 and to the tip of the short arm of the Y chromosome (Yp). In our patient this locus is present both in Xq21 and in Xq28, in addition to the Yp, being thus duplicated in the X chromosome. Sequence analysis of the 3' breakpoint suggested that an illegitimate recombination is probably the cause of this complex rearrangement.

Adult↗
PubMed1994

X-linked spastic paraplegia (SPG1), MASA syndrome and X-linked hydrocephalus result from mutations in the L1 gene.

X-linked hydrocephalus, spastic paraplegia type I and MASA syndrome are related disorders with loci in subchromosomal region Xq28. We have previously shown that X-linked hydrocephalus is caused by mutations in the gene for neural cell adhesion molecule L1 (L1CAM), an axonal glycoprotein involved in neuronal migration and differentiation. Here we report mutations of the L1 gene in MASA syndrome and SPG1, in addition to HSAS families. Two of the HSAS mutations would abolish cell surface expression of L1 and represent the first functional null mutations in this disorder. Our results indicate that these three syndromes from part of a clinical spectrum resulting from a heterogeneous group of mutations in the L1 gene.

Aphasia↗