Phosphorylation of spore coat proteins by a family of atypical protein kinases

Nguyen, Kim B.; Sreelatha, Anju; Durrant, Eric; López‐Garrido, Javier; Muszewska, Anna; Dudkiewicz, Małgorzata; Grynberg, Marcin; Yee, Samantha S.; Pogliano, Kit; Tomchick, Diana R.; Pawłowski, Krzysztof; Dixon, Jack E.; Tagliabracci, Vincent S.

doi:10.1073/pnas.1605917113

Cited by 58 publications

(88 citation statements)

References 78 publications

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“…E. coli SelO was cloned without a tag into the MCS2 of petDuet1. SelO D256A, sucA T405A and sucA S404A were generated by site directed mutagenesis as described (Nguyen et al, 2016). Briefly, primers were designed using the Agilent Quick Change Primer design tool: https://www.genomics.agilent.com/primerDesignProgram.jsp and used in PCR reaction to generate the desired mutation using PfuTurbo DNA polymerase (Thermo).…”

Section: Star Methods Textmentioning

confidence: 99%

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Sreelatha

Yee

López

et al. 2018

Cell

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Summary Approximately 10% of human protein kinases are believed to be inactive and named pseudokinases because they lack residues required for catalysis. Here we show that the highly conserved pseudokinase selenoprotein-O (SelO) transfers AMP from ATP to Ser, Thr and Tyr residues on protein substrates (AMPylation), uncovering a previously unrecognized activity for a member of the protein kinase superfamily. The crystal structure of a SelO homolog reveals a protein kinase-like fold with ATP flipped in the active site, thus providing a structural basis for catalysis. SelO pseudokinases localize to the mitochondria and AMPylate proteins involved in redox homeostasis. Consequently, SelO activity is necessary for the proper cellular response to oxidative stress. Our results suggest that AMPylation may be a more widespread post translational modification than previously appreciated and that pseudokinases should be analyzed for alternative transferase activities.

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Section: Star Methods Textmentioning

confidence: 99%

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Sreelatha

Yee

López

et al. 2018

Cell

Self Cite

162

224

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“…The second accessory gene group consisted of a contiguous set of genes whose protein products are similar to the kinase CotH [35], a poly-phosphate polymerase, and two conserved domains of unknown function, DUF4956 and DUF4832, that are contiguous with the pelDEA DA FG gene cluster in Streptococcal and Clostridial species (Fig. S4).…”

Section: Resultsmentioning

confidence: 99%

Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene cluster

Whitfield

Marmont

Bundalovic-Torma

et al. 2019

Preprint

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AbstractOur understanding of the biofilm matrix components utilized by Gram-positive bacteria, and the signalling pathways that regulate their production are largely unknown. In a companion study, we developed a computational pipeline for the unbiased identification of homologous bacterial operons and applied this algorithm to the analysis of synthase-dependent exopolysaccharide biosynthetic systems (https://doi.org/10.1101/769745). Here, we explore the finding that many species of Gram-positive bacteria have operons with similarity to the Pseudomonas aeruginosa pel locus. Our characterization of the pelDEADAFG operon from Bacillus cereus ATCC 10987, presented herein, demonstrates that this locus is required for biofilm formation and produces a polysaccharide structurally similar to Pel. We show that the degenerate GGDEF domain of the B. cereus PelD ortholog binds cyclic-3’,5’-dimeric guanosine monophosphate (c-di-GMP), and that this binding is required for biofilm formation. Finally, we identify a diguanylate cyclase, CdgF, and a c-di-GMP phosphodiesterase, CdgE, that reciprocally regulate the production of Pel. The discovery of this novel c-di-GMP regulatory circuit significantly contributes to our limited understanding of c-di-GMP signalling in Gram-positive organisms. Furthermore, conservation of the core pelDEADAFG locus amongst many species of Bacilli, Clostridia, Streptococci, and Actinobacteria suggests that Pel may be a common biofilm matrix component in many Gram-positive bacteria.Author summaryThe Pel polysaccharide is required for biofilm formation in P. aeruginosa and we have previously found that the genes necessary for biosynthesis of this polymer are broadly distributed across Gram-negative bacteria. Herein, we show that many species of Gram-positive bacteria also possess Pel biosynthetic genes and demonstrate that these genes are used Bacillus cereus for biofilm formation. We show that Pel production in B. cereus is regulated by c-di-GMP and have identified two enzymes, a diguanylate cyclase, CdgF, and a phosphodiesterase, CdgE, that control the levels of this bacterial signalling molecule. While Pel production in B. cereus also requires the binding of c-di-GMP to the receptor PelD, the divergence of this protein in Streptococci suggests a c-di-GMP independent mechanism of regulation is used in this species. The discovery of a Pel biosynthetic gene cluster in Gram-positive bacteria and our characterization of the pel operon in B. cereus suggests that Pel is a widespread biofilm component across all bacteria.

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“…CotH are atypical protein kinases that constitute a structural component of spores of different human pathogens, including prokaryotes and eukaryotes [95,96]. In Bacillus subtilis, CotH regulates the integrity of the spore, as CotH mutant spores showed a deficiency in germination [97].…”

Section: Coth Gene Family a Distinctive Virulence Factor In Mucoralesmentioning

confidence: 99%

Genes, Pathways, and Mechanisms Involved in the Virulence of Mucorales

Lax

Pérez-Arques

Navarro-Mendoza

et al. 2020

Genes

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The order Mucorales is a group of ancient fungi with limited tools for gene manipulation. The main consequence of this manipulation unwillingness is the limited knowledge about its biology compared to other fungal groups. However, the emerging of mucormycosis, a fungal infection caused by Mucorales, is attracting the medical spotlight in recent years because the treatments available are not efficient in reducing the high mortality associated with this disease. The result of this renewed interest in Mucorales and mucormycosis is an extraordinarily productive effort to unveil their secrets during the last decade. In this review, we describe the most compelling advances related to the genetic study of virulence factors, pathways, and molecular mechanisms developed in these years. The use of a few genetic study models has allowed the characterization of virulence factors in Mucorales that were previously described in other pathogens, such as the uptake iron systems, the mechanisms of dimorphism, and azole resistances. More importantly, recent studies are identifying new genes and mechanisms controlling the pathogenic potential of Mucorales and their interactions with the host, offering new alternatives to develop specific strategies against mucormycosis.Genes 2020, 11, 317 2 of 17 Mucorales are a neglected phylogenetic group compared to others such as Ascomycetes and Basidiomycetes. The limited knowledge about the genetics of Mucorales is a consequence of the restricted tools for gene manipulation, as most of them cannot be transformed. However, DNA can be introduced in Mucor circinelloides, Rhizopus delemar, and Rhizopus oryzae [6,7]. These genetic models and the alarm raised for the emerging cases of mucormycosis are attracting the interest of the scientific community. Thus, the last decade has produced several studies related to genes, pathways, and mechanisms showing a direct connection with virulence in Mucorales [8,9]. One of the most studied mechanisms has been the process of gene silencing or RNA interfering (RNAi) in M. circinelloides [10]. After the dissection of the gene silencing machinery, the knowledge of this mechanism allowed the unveiling of a new and particular type of antifungal resistance mediated by temporal epigenetic changes [11]. In addition, the applied use of gene silencing enabled the development of functional genomics techniques, which have been used for the identification of several new virulence factors [12]. Along with silencing, gene disruption driven by homologous recombination has also allowed the study of the particular role in M. circinelloides of virulence factors identified in other fungi, such as the role of a high-affinity iron uptake mechanism, the protein family of CotH, and the calcineurin pathway. Moreover, the implementation of the new omics technologies has produced a long list of candidate genes not previously related to virulence, becoming promising targets for the development of new treatments against mucormycosis. Finally, the diversity of molecular and cell methodolo...

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Phosphorylation of spore coat proteins by a family of atypical protein kinases

Cited by 58 publications

References 78 publications

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene cluster

Genes, Pathways, and Mechanisms Involved in the Virulence of Mucorales

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