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Full-Text Articles in Molecular Biology

Orbit: A New Paradigm For Genetic Engineering Of Mycobacterial Chromosomes, Kenan C. Murphy, Samantha J. Nelson, Subhalaxmi Nambi, Kadamba Papavinasasundaram, Christina E. Baer, Christopher M. Sassetti Dec 2018

Orbit: A New Paradigm For Genetic Engineering Of Mycobacterial Chromosomes, Kenan C. Murphy, Samantha J. Nelson, Subhalaxmi Nambi, Kadamba Papavinasasundaram, Christina E. Baer, Christopher M. Sassetti

Open Access Articles

Two efficient recombination systems were combined to produce a versatile method for chromosomal engineering that obviates the need to prepare double-stranded DNA (dsDNA) recombination substrates. A synthetic "targeting oligonucleotide" is incorporated into the chromosome via homologous recombination mediated by the phage Che9c RecT annealase. This oligonucleotide contains a site-specific recombination site for the directional Bxb1 integrase (Int), which allows the simultaneous integration of a "payload plasmid" that contains a cognate recombination site and a selectable marker. The targeting oligonucleotide and payload plasmid are cotransformed into a RecT- and Int-expressing strain, and drug-resistant homologous recombinants are selected in a single step ...


The Intra-S Checkpoint Responses To Dna Damage, Divya Ramalingam Iyer, Nicholas R. Rhind Feb 2017

The Intra-S Checkpoint Responses To Dna Damage, Divya Ramalingam Iyer, Nicholas R. Rhind

Open Access Articles

Faithful duplication of the genome is a challenge because DNA is susceptible to damage by a number of intrinsic and extrinsic genotoxins, such as free radicals and UV light. Cells activate the intra-S checkpoint in response to damage during S phase to protect genomic integrity and ensure replication fidelity. The checkpoint prevents genomic instability mainly by regulating origin firing, fork progression, and transcription of G1/S genes in response to DNA damage. Several studies hint that regulation of forks is perhaps the most critical function of the intra-S checkpoint. However, the exact role of the checkpoint at replication forks has ...


A Gene-Centered C. Elegans Protein-Dna Interaction Network Provides A Framework For Functional Predictions, Juan Fuxman Bass, Carles Pons, Lucie Kozlowski, John S. Reece-Hoyes, Shaleen Shrestha, Amy D. Holdorf, Akihiro Mori, Chad L. Myers, Albertha J. M. Walhout Oct 2016

A Gene-Centered C. Elegans Protein-Dna Interaction Network Provides A Framework For Functional Predictions, Juan Fuxman Bass, Carles Pons, Lucie Kozlowski, John S. Reece-Hoyes, Shaleen Shrestha, Amy D. Holdorf, Akihiro Mori, Chad L. Myers, Albertha J. M. Walhout

Open Access Articles

Transcription factors (TFs) play a central role in controlling spatiotemporal gene expression and the response to environmental cues. A comprehensive understanding of gene regulation requires integrating physical protein-DNA interactions (PDIs) with TF regulatory activity, expression patterns, and phenotypic data. Although great progress has been made in mapping PDIs using chromatin immunoprecipitation, these studies have only characterized ~10% of TFs in any metazoan species. The nematode C. elegans has been widely used to study gene regulation due to its compact genome with short regulatory sequences. Here, we delineated the largest gene-centered metazoan PDI network to date by examining interactions between 90 ...


The Molecular Basis Of The Genesis Of Basal Tone In Internal Anal Sphincter, Cheng-Hai Zhang, Donghai Lui, Ping Lu, Kaizhi Zheng, Siobhan M. Craige, Lawrence Lifshitz, John F. Keaney Jr., Kevin E. Fogarty, Ronghua Zhuge, Min-Sheng Zhu Apr 2016

The Molecular Basis Of The Genesis Of Basal Tone In Internal Anal Sphincter, Cheng-Hai Zhang, Donghai Lui, Ping Lu, Kaizhi Zheng, Siobhan M. Craige, Lawrence Lifshitz, John F. Keaney Jr., Kevin E. Fogarty, Ronghua Zhuge, Min-Sheng Zhu

Open Access Articles

Smooth muscle sphincters exhibit basal tone and control passage of contents through organs such as the gastrointestinal tract; loss of this tone leads to disorders such as faecal incontinence. However, the molecular mechanisms underlying this tone remain unknown. Here, we show that deletion of myosin light-chain kinases (MLCK) in the smooth muscle cells from internal anal sphincter (IAS-SMCs) abolishes basal tone, impairing defecation. Pharmacological regulation of ryanodine receptors (RyRs), L-type voltage-dependent Ca(2+) channels (VDCCs) or TMEM16A Ca(2+)-activated Cl(-) channels significantly changes global cytosolic Ca(2+) concentration ([Ca(2+)]i) and the tone. TMEM16A deletion in IAS-SMCs abolishes ...


Comparative Genomics Reveals Chd1 As A Determinant Of Nucleosome Spacing In Vivo, Amanda Hughes, Oliver J. Rando Jul 2015

Comparative Genomics Reveals Chd1 As A Determinant Of Nucleosome Spacing In Vivo, Amanda Hughes, Oliver J. Rando

Open Access Articles

Packaging of genomic DNA into nucleosomes is nearly universally conserved in eukaryotes, and many features of the nucleosome landscape are quite conserved. Nonetheless, quantitative aspects of nucleosome packaging differ between species because, for example, the average length of linker DNA between nucleosomes can differ significantly even between closely related species. We recently showed that the difference in nucleosome spacing between two Hemiascomycete species-Saccharomyces cerevisiae and Kluyveromyces lactis-is established by trans-acting factors rather than being encoded in cis in the DNA sequence. Here, we generated several S. cerevisiae strains in which endogenous copies of candidate nucleosome spacing factors are deleted and ...


Single Point Mutation In Rabenosyn-5 In A Female With Intractable Seizures And Evidence Of Defective Endocytotic Trafficking, Sylvia Stockler, Silvia Corvera, David G. Lambright, Kevin E. Fogarty, Ekaterina Nosova, Deborah Marie Leonard, Robert Steinfeld, Cameron Ackerley, Casper Shyr, Nicolas Au, Kathrin Selby, Margot Van Allen, Hilary Vallance, Ron Wevers, David Watkins, David Rosenblatt, Colin J. Ross, Elizabeth Conibear, Wyeth Wasserman, Clara Van Karnebeek Sep 2014

Single Point Mutation In Rabenosyn-5 In A Female With Intractable Seizures And Evidence Of Defective Endocytotic Trafficking, Sylvia Stockler, Silvia Corvera, David G. Lambright, Kevin E. Fogarty, Ekaterina Nosova, Deborah Marie Leonard, Robert Steinfeld, Cameron Ackerley, Casper Shyr, Nicolas Au, Kathrin Selby, Margot Van Allen, Hilary Vallance, Ron Wevers, David Watkins, David Rosenblatt, Colin J. Ross, Elizabeth Conibear, Wyeth Wasserman, Clara Van Karnebeek

Open Access Articles

Background. We report a 6.5 year-old female with a homozygous missense mutation in ZFYVE20, encoding Rabenosyn-5 (Rbsn-5), a highly conserved multi-domain protein implicated in receptor-mediated endocytosis. The clinical presentation includes intractable seizures, developmental delay, microcephaly, dysostosis, osteopenia, craniofacial dysmorphism, macrocytosis and megaloblastoid erythropoiesis. Biochemical findings include transient cobalamin deficiency, severe hypertriglyceridemia upon ketogenic diet, microalbuminuria and partial cathepsin D deficiency.

Methods and results. Whole exome sequencing followed by Sanger sequencing confirmed a rare (frequency:0.003987) homozygous missense mutation, g.15,116,371 G inverted question mark > inverted question markA (c.1273G inverted question mark > inverted question markA), in ...