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1. Competence pili in Streptococcus pneumoniae are highly dynamic structures that retract to promote DNA uptake

   https://doi.org/10.1111/mmi.14718  

   Lam, Trinh ; Ellison, Courtney K. ; Eddington, David T. ; Brun, Yves V. ; Dalia, Ankur B. ; Morrison, Donald A. ( April 2021 , Molecular Microbiology)

   The competence pili of transformable Gram‐positive species are phylogenetically related to the diverse and widespread class of extracellular filamentous organelles known as type IV pili. In Gram‐negative bacteria, type IV pili act through dynamic cycles of extension and retraction to carry out diverse activities including attachment, motility, protein secretion, and DNA uptake. It remains unclear whether competence pili in Gram‐positive species exhibit similar dynamic activity, and their mechanism of action for DNA uptake remains unclear. They are hypothesized to either (1) leave transient cavities in the cell wall that facilitate DNA passage, (2) form static adhesins to enrich DNA near the cell surface for subsequent uptake by membrane‐embedded transporters, or (3) play an active role in translocating bound DNA via dynamic activity. Here, we use a recently described pilus labeling approach to demonstrate that competence pili inStreptococcus pneumoniaeare highly dynamic structures that rapidly extend and retract from the cell surface. By labeling the principal pilus monomer, ComGC, with bulky adducts, we further demonstrate that pilus retraction is essential for natural transformation. Together, our results suggest that Gram‐positive competence pili in other species may also be dynamic and retractile structures that play an active role in DNA uptake.

    

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2. Random transposon mutagenesis identifies genes essential for transformation in Methanococcus maripaludis

   https://doi.org/10.1007/s00438-023-01994-7  

   Fonseca, Dallas R. ; Loppnow, Madison B. ; Day, Leslie A. ; Kelsey, Elisa L. ; Abdul Halim, Mohd Farid ; Costa, Kyle C. ( February 2023 , Molecular Genetics and Genomics)

   Natural transformation, the process whereby a cell acquires DNA directly from the environment, is an important driver of evolution in microbial populations, yet the mechanism of DNA uptake is only characterized in bacteria. To expand our understanding of natural transformation in archaea, we undertook a genetic approach to identify a catalog of genes necessary for transformation inMethanococcus maripaludis. Using an optimized method to generate random transposon mutants, we screened 6144 mutant strains for defects in natural transformation and identified 25 transformation-associated candidate genes. Among these are genes encoding components of the type IV-like pilus, transcription/translation associated genes, genes encoding putative membrane bound transport proteins, and genes of unknown function. Interestingly, similar genes were identified regardless of whether replicating or integrating plasmids were provided as a substrate for transformation. Using allelic replacement mutagenesis, we confirmed that several genes identified in these screens are essential for transformation. Finally, we identified a homolog of a membrane bound substrate transporter inMethanoculleus thermophilusand verified its importance for transformation using allelic replacement mutagenesis, suggesting a conserved mechanism for DNA transfer in multiple archaea. These data represent an initial characterization of the genes important for transformation which will inform efforts to understand gene flow in natural populations. Additionally, knowledge of the genes necessary for natural transformation may assist in identifying signatures of transformation machinery in archaeal genomes and aid the establishment of new model genetic systems for studying archaea.

    

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3. Overexpression of VIRE2-INTERACTING PROTEIN2 in Arabidopsis regulates genes involved in Agrobacterium-mediated plant transformation and abiotic stresses

   https://doi.org/10.1038/s41598-019-49590-3  

   Raman, Vidhyavathi ; Anand, Ajith ; Vasudevan, Balaji ; Morsy, Mustafa R. ; Pant, Bikram D. ; Lee, Hee-Kyung ; Tang, Yuhong ; Mysore, Kirankumar S. ( September 2019 , Scientific Reports)

   ArabidopsisVIRE2-INTERACTINGPROTEIN2 (VIP2) was previously described as a protein with a NOT domain, and Arabidopsisvip2mutants are recalcitrant toAgrobacterium-mediated root transformation. Here we show that VIP2 is a transcription regulator and the C-terminal NOT2 domain of VIP2 interacts with VirE2. Interestingly,AtVIP2overexpressor lines in Arabidopsis did not show an improvement inAgrobacterium-mediated stable root transformation, but the transcriptome analysis identified 1,634 differentially expressed genes compared to wild-type. These differentially expressed genes belonged to various functional categories such as membrane proteins, circadian rhythm, signaling, response to stimulus, regulation of plant hypersensitive response, sequence-specific DNA binding transcription factor activity and transcription regulatory region binding. In addition to regulating genes involved inAgrobacterium-mediated plant transformation,AtVIP2overexpressor line showed differential expression of genes involved in abiotic stresses. The majority of the genes involved in abscisic acid (ABA) response pathway, containing the Abscisic Acid Responsive Element (ABRE) element within their promoters, were down-regulated inAtVIP2overexpressor lines. Consistent with this observation,AtVIP2overexpressor lines were more susceptible to ABA and other abiotic stresses. Based on the above findings, we hypothesize that VIP2 not only plays a role inAgrobacterium-mediated plant transformation but also acts as a general transcriptional regulator in plants.

    

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4. Harnessing Multiple, Nonproteogenic Substitutions to Optimize CSP:ComD Hydrophobic Interactions in Group 1 Streptococcus pneumoniae

   https://doi.org/10.1002/cbic.202000876  

   Milly, Tahmina A. ; Engler, Emilee R. ; Chichura, Kylie S. ; Buttner, Alec R. ; Koirala, Bimal ; Tal‐Gan, Yftah ; Bertucci, Michael A. ( April 2021 , ChemBioChem)

   Streptococcus pneumoniae(pneumococcus) is a human pathobiont that causes drastic antibiotic‐resistant infections and is responsible for millions of deaths universally. Pneumococcus pathogenicity relies on the competence‐stimulating peptide (CSP)‐mediated quorum‐sensing (QS) pathway that controls competence development for genetic transformation and, consequently, the spread of antibiotic resistance and virulence genes. Modulation of QS inS. pneumoniaecan therefore be used to enervate pneumococcal infectivity as well as minimize the susceptibility to resistance development. In this work, we sought to optimize the interaction of CSP1 with its cognate transmembrane histidine kinase receptor (ComD1) through substitution of proteogenic and nonproteogenic amino acids on the hydrophobic binding face of CSP1. The findings from this study not only provided additional structure–activity data that are significant in optimizing CSP1 potency, but also led to the development of potent QS modulators. These CSP‐based QS modulators could be used as privileged scaffolds for the development of antimicrobial agents against pneumococcal infections.

    

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5. Phototaxis in a wild isolate of the cyanobacterium Synechococcus elongatus

   https://doi.org/10.1073/pnas.1812871115  

   Yang, Yiling ; Lam, Vinson ; Adomako, Marie ; Simkovsky, Ryan ; Jakob, Annik ; Rockwell, Nathan C. ; Cohen, Susan E. ; Taton, Arnaud ; Wang, Jingtong ; Lagarias, J. Clark ; et al ( December 2018 , Proceedings of the National Academy of Sciences)

   Many cyanobacteria, which use light as an energy source via photosynthesis, have evolved the ability to guide their movement toward or away from a light source. This process, termed “phototaxis,” enables organisms to localize in optimal light environments for improved growth and fitness. Mechanisms of phototaxis have been studied in the coccoid cyanobacteriumSynechocystissp. strain PCC 6803, but the rod-shapedSynechococcus elongatusPCC 7942, studied for circadian rhythms and metabolic engineering, has no phototactic motility. In this study we report a recent environmental isolate ofS. elongatus, the strain UTEX 3055, whose genome is 98.5% identical to that of PCC 7942 but which is motile and phototactic. A six-gene operon encoding chemotaxis-like proteins was confirmed to be involved in phototaxis. Environmental light signals are perceived by a cyanobacteriochrome, PixJ_Se(Synpcc7942_0858), which carries five GAF domains that are responsive to blue/green light and resemble those of PixJ fromSynechocystis. Plate-based phototaxis assays indicate that UTEX 3055 uses PixJ_Seto sense blue and green light. Mutation of conserved functional cysteine residues in different GAF domains indicates that PixJ_Secontrols both positive and negative phototaxis, in contrast to the multiple proteins that are employed for implementing bidirectional phototaxis inSynechocystis.

    

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