Si Lok

Bio: Si Lok is an academic researcher from University of Toronto. The author has contributed to research in topics: Regulation of gene expression & Gene family. The author has an hindex of 5, co-authored 5 publications receiving 6753 citations.

Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA.

Abstract: Overlapping complementary DNA clones were isolated from epithelial cell libraries with a genomic DNA segment containing a portion of the putative cystic fibrosis (CF) locus, which is on chromosome 7 Transcripts, approximately 6500 nucleotides in size, were detectable in the tissues affected in patients with CF The predicted protein consists of two similar motifs, each with (i) a domain having properties consistent with membrane association and (ii) a domain believed to be involved in ATP (adenosine triphosphate) binding A deletion of three base pairs that results in the omission of a phenylalanine residue at the center of the first predicted nucleotide-binding domain was detected in CF patients

Gamma-crystallin family of the mouse lens: structural and evolutionary relationships

Abstract: The heterogeneity inherent among gamma-crystallins of the mouse lens was investigated by sequence analysis of three gamma-crystallin-specific cDNAs. Comparison of the nucleotide sequence of these cDNAs and one previously reported by us revealed that the four gamma-cDNAs share 80-90% homology in nucleotide sequence. The entire 3' half of the coding region shows more variability than the 5' half, whereas the greatest variability is observed in the 3' untranslated region where numerous base substitutions, deletions, and insertions seem to have occurred. Alignment of the amino acid sequences of the four mouse gamma-crystallins according to the known four structural motifs of the major calf gamma-crystallin, gamma-II, suggests that all four mouse polypeptides are structurally very similar to calf gamma-II. However, most of the mouse polypeptides differ from gamma-II by the absence of one amino acid residue, resulting in a shorter connecting peptide between the two globular domains of the protein. Primary sequence alignment also revealed that the four mouse gamma-crystallins are most divergent in the third structural motif of the polypeptide. The significance of these differences in terms of the structure and function of the gamma-crystallins in the mouse lens is discussed.

Lens-specific promoter activity of a mouse gamma-crystallin gene.

Abstract: Crystallins are the major water-soluble proteins in vertebrate eye lenses. These lens-specific proteins are encoded by several gene families, and their expression is differentially regulated during lens cell differentiation. Here we show that a cloned mouse gamma-crystallin promoter is active in lens explants derived from 14-day-old chicken embryos but inactive in a variety of cells of non-lens origin. We also show that sequences required for proper utilization of this promoter are contained between nucleotide positions -392 and +47 relative to the transcription initiation site; deletion of sequences from positions -392 to -171 completely abolishes promoter activity. Since chickens do not have gamma-crystallin genes, the expression of a mouse gamma-crystallin promoter in chicken lens cells suggests that different classes of crystallin genes may be regulated by common lens tissue-specific mechanism(s) independent of species.

Analysis of the mouse γ-crystallin gene family: assignment of multiple cDNAs to discrete genomic sequences and characterization of a representative gene

Abstract: Blot hybridization analysis of mouse DNA with gamma-crystallin-specific cDNAs has detected the presence of a multigene family comprised of at least four related genes. The detailed structure of one of these genes, mouse gamma 4-crystallin (M gamma 4.1), and its corresponding cDNA has been determined. The gene spans approximately 2.6 kilobases (kb) and contains two introns. The gene predicts a polypeptide of 174 amino acids that shares extensive sequence homology with gamma-crystallin polypeptides of other species. The two similar structural domains of the protein correspond exactly to the second and third exons of the gene, supporting an exon-duplication model of gene evolution. The similarity in structure of this gene to that recently reported for a gamma-crystallin gene of the rat (1) suggests that a common structure may exist for all gamma-crystallin genes of the two species. Moreover, a highly conserved region, 50 nucleotides in length, immediately precedes the TATA box of both the mouse and rat genes, suggesting that this sequence may be important in gene regulation.

ABC Transporters: From Microorganisms to Man

Abstract: INTRODUCTION .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67 DOMAIN ORGANIZATION: The Typical ABC Transporter . . . . . . . . . . . . . . . . . 73 THE TRANSMEMBRANE DOMAINS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76 The "Two-Times-Six" Helix Paradigm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76 Sequence Similarities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78 THE ATP-BINDING DOMAINS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79 PERIPLASMIC-BINDING PROTEINS ... . 84 SUBSTRATE SPECIFICITY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86 THE ROLE OF ATP : Coupling Energy to Transport . . . . . . . . . . . . . . . . . . . " . . . . . 88 COVALENT MODIFICATION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91 CELLULAR FUNCTIONS OF ABC TRANSPORTERS . . . . . . . . . . . . . . . . . . . . . . . 9 1 Nutrient Uptake . . . . . . . . . . . . ....... . 9 1 Protein Export ....... . .... . . . . . . . . . . . . ... . ......... . ...... . ... . .. 93 Intracellular Membranes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93 Regulation of ABC Transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94 Regulation by ABC Transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95 Drug and Antibiotic Resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95 Channel Functions: CFTR and P-glycoprotein . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97 MECHANISMS OF SOLUTE TRANSLOCATION . . . . . . . . . . . . . . . . . . . . . . . . . . 98 Structure of the Transmembrane Complex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99 Channels and Transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 0 I Energy Coupling andlor Gating . 102 CONCLUDING REMARKS . 103

Identification of the cystic fibrosis gene: genetic analysis.

Abstract: Approximately 70 percent of the mutations in cystic fibrosis patients correspond to a specific deletion of three base pairs, which results in the loss of a phenylalanine residue at amino acid position 508 of the putative product of the cystic fibrosis gene. Extended haplotype data based on DNA markers closely linked to the putative disease gene locus suggest that the remainder of the cystic fibrosis mutant gene pool consists of multiple, different mutations. A small set of these latter mutant alleles (about 8 percent) may confer residual pancreatic exocrine function in a subgroup of patients who are pancreatic sufficient. The ability to detect mutations in the cystic fibrosis gene at the DNA level has important implications for genetic diagnosis.

Identification of the cystic fibrosis gene: Chromosome walking and jumping

Abstract: An understanding of the basic defect in the inherited disorder cystic fibrosis requires cloning of the cystic fibrosis gene and definition of its protein product. In the absence of direct functional information, chromosomal map position is a guide for locating the gene. Chromosome walking and jumping and complementary DNA hybridization were used to isolate DNA sequences, encompassing more than 500,000 base pairs, from the cystic fibrosis region on the long arm of human chromosome 7. Several transcribed sequences and conserved segments were identified in this cloned region. One of these corresponds to the cystic fibrosis gene and spans approximately 250,000 base pairs of genomic DNA.

Overexpression of a transporter gene in a multidrug-resistant human lung cancer cell line

Abstract: The doxorubicin-selected lung cancer cell line H69AR is resistant to many chemotherapeutic agents. However, like most tumor samples from individuals with this disease, it does not overexpress P-glycoprotein, a transmembrane transport protein that is dependent on adenosine triphosphate (ATP) and is associated with multidrug resistance. Complementary DNA (cDNA) clones corresponding to messenger RNAs (mRNAs) overexpressed in H69AR cells were isolated. One cDNA hybridized to an mRNA of 7.8 to 8.2 kilobases that was 100- to 200-fold more expressed in H69AR cells relative to drug-sensitive parental H69 cells. Overexpression was associated with amplification of the cognate gene located on chromosome 16 at band p13.1. Reversion to drug sensitivity was associated with loss of gene amplification and a marked decrease in mRNA expression. The mRNA encodes a member of the ATP-binding cassette transmembrane transporter superfamily.

Genome-wide genetic marker discovery and genotyping using next-generation sequencing.

Abstract: The authors describe the best practices for a growing number of methods that use next-generation sequencing to rapidly discover and assess genetic markers across any genome, with applications from population genomics and quantitative trait locus mapping to marker-assisted selection.