High-resolution landscape of an antibiotic binding site.

Antibiotic binding sites are located in important domains of essential enzymes and have been extensively studied in the context of resistance mutations; however, their study is limited by positive selection. Using multiplex genome engineering to overcome this constraint, we generate and characterize a collection of 760 single-residue mutants encompassing the entire rifampicin binding site of Escherichia coli RNA polymerase (RNAP). By genetically mapping drug-enzyme interactions, we identify an alpha helix where mutations considerably enhance or disrupt rifampicin binding.

MRLC controls apoptotic cell death and functions to regulate epidermal development during planarian regeneration and homeostasis.

Adult stem cells (ASCs) are pluripotent cells with the capacity to self-renew and constantly replace lost cells due to physiological turnover or injury. Understanding the molecular mechanisms of the precise coordination of stem cell proliferation and proper cell fate decision is important to regeneration and organismal homeostasis. The planarian epidermis provides a highly tractable model to study ASC complex dynamic due to the distinct spatiotemporal differentiation stages during lineage development.

RNA polymerase drives ribonucleotide excision DNA repair in E. coli.

Ribonuclease HII (RNaseHII) is the principal enzyme that removes misincorporated ribonucleoside monophosphates (rNMPs) from genomic DNA. Here, we present structural, biochemical, and genetic evidence demonstrating that ribonucleotide excision repair (RER) is directly coupled to transcription. Affinity pull-downs and mass-spectrometry-assisted mapping of in cellulo inter-protein cross-linking reveal the majority of RNaseHII molecules interacting with RNA polymerase (RNAP) in E.

Rho-dependent transcription termination: a revisionist view.

Rho is a hexameric bacterial RNA helicase, which became a paradigm of factor-dependent transcription termination. The broadly accepted ("textbook") model posits a series of steps, wherein Rho first binds C-rich () sites on nascent RNA, uses its ATP-dependent translocase activity to catch up with RNA polymerase (RNAP), and either pulls the transcript from the elongation complex or pushes RNAP forward, thus terminating transcription. However, this appealingly simple mechano-chemical model lacks a biological realism and is increasingly at odds with genetic and biochemical data.

Analysing the fitness cost of antibiotic resistance to identify targets for combination antimicrobials.

Mutations in the rifampicin (Rif)-binding site of RNA polymerase (RNAP) confer antibiotic resistance and often have global effects on transcription that compromise fitness and stress tolerance of resistant mutants. We suggested that the non-essential genome, through its impact on the bacterial transcription cycle, may represent an untapped source of targets for combination antimicrobial therapies. Using transposon sequencing, we carried out a genome-wide analysis of fitness cost in a clinically common rpoB H526Y mutant.

TBX2/3 is required for regeneration of dorsal-ventral and medial-lateral polarity in planarians.

The molecular mechanisms responsible for axis establishment during non-embryonic processes remain elusive. The planarian flatworm is an ideal model organism to study body axis polarization and patterning in vivo. Here, we identified a homolog of the TBX2/3 in the planarian Dugesia japonica.

Tubgcp3 is a mitotic regulator of planarian epidermal differentiation.

Tubgcp3/GCP3 (The centrosomal protein γ-tubulin complex protein 3) is a component of the γ-tubulin small complexes (γ-TuSCs) and γ-tubulin ring complexes (γ-TuRCs), which play critical roles in mitotic spindle formation during mitosis. However, its function in stem cell development has not been thoroughly elucidated. The planarian flatworm, which contains a large number of adult somatic stem cells (neoblasts), is a unique model to study stem cell lineage development in vivo.

Pre-termination Transcription Complex: Structure and Function.

Rho is a general transcription termination factor playing essential roles in RNA polymerase (RNAP) recycling, gene regulation, and genomic stability in most bacteria. Traditional models of transcription termination postulate that hexameric Rho loads onto RNA prior to contacting RNAP and then translocates along the transcript in pursuit of the moving RNAP to pull RNA from it. Here, we report the cryoelectron microscopy (cryo-EM) structures of two termination process intermediates.

Cdc42 regulate the apoptotic cell death required for planarian epidermal regeneration and homeostasis.

Rho GTPases have been shown previously to play important roles in several cellular processes by regulating the organization of the actin and microtubule cytoskeletons. However, the mechanisms of Rho GTPases that integrate the cellular responses during regeneration have not been thoroughly elucidated. The planarian flatworm, which contains a large number of adult somatic stem cells (neoblasts), is a unique model to study stem cell lineage development in vivo.

Structure of human endonuclease V as an inosine-specific ribonuclease.

The 6-aminopurine ring of adenosine (A) can be deaminated to form the 6-oxopurine of inosine (I). Endonuclease Vs (EndoVs) are inosine-specific nucleases that cleave at the second phosphodiester bond 3' to inosine. EndoV proteins are highly conserved in all domains of life, but the bacterial and human enzymes seem to display distinct substrate preferences.

Cocrystal structures of glycyl-tRNA synthetase in complex with tRNA suggest multiple conformational states in glycylation.

Aminoacyl-tRNA synthetases are an ancient enzyme family that specifically charges tRNA molecules with cognate amino acids for protein synthesis. Glycyl-tRNA synthetase (GlyRS) is one of the most intriguing aminoacyl-tRNA synthetases due to its divergent quaternary structure and abnormal charging properties. In the past decade, mutations of human GlyRS (hGlyRS) were also found to be associated with Charcot-Marie-Tooth disease.