Biology subjects

Pratte, B.

Publications and source records attributed to Pratte, B..

3 recordsLinked to original sources

bioRxiv2025

Excision of a 37-kb excision element as a circular plasmid in the cyanobacterium Anabaena variabilis

Anabaena (aka Trichormus) variabilis ATCC 29413 strain FD is a filamentous, heterocyst-forming cyanobacterium with a 6.36 Mb chromosome, three circular plasmids, A (366 kb), B (35.8 kb), C (301 kb), and a 37-kb excision element. This 37-kb element in A. variabilis ATCC 29413 strain FD was integrated into the tRNAcys gene but was absent in the very closely related strains A. variabilis FSR and PNB. The 37-kb element was also excised as a circular molecule at a very low frequency compared to the integrated form. The 37-kb element has a partial copy of tRNAcys; therefore, integration produced a functional but genetically distinct copy of tRNAcys. Integration and excision are likely mediated by a putative integrase with a tyrosine recombinase domain encoded within the element, possibly using the duplicated copies of the 7-bp anticodon of the tRNAcys as a recombination site. Like many other bacterial elements, the function of this element is unknown, but, like other well-characterized excision elements in heterocystous cyanobacteria, it does not appear to be important for survival.

genetics↗

bioRxiv2025

A plasmid-encoded type IIB restriction-modification system in cyanobacteria blocks conjugative gene transfer

Anabaena (aka Trichormus) variabilis ATCC 29413 is a filamentous, heterocyst-forming cyanobacterium with a 6.36 Mb chromosome, four circular plasmids, A (366 kb), B (35.8 kb), C (301 kb), and D (27 kb), and a 37-kb excision element. The hsdRMS genes on plasmid D may encode a type IIB restriction-modification system that protects the cells from invasion by foreign DNA. A variant of A. variabilis, strain FD, lacking plasmid D, grew with the same generation time as A. variabilis ATCC 29413, which stably maintained plasmid D. Nostoc sp. PCC 7120 and Nostoc sp. M131, which lack plasmid D and hsdRMS, grew similarly with a synthetic replicating plasmid, with or without added hsdRMS genes. Although the natural plasmid D was very stable in A. variabilis ATCC 29413, the synthetic plasmid was easily lost in Nostoc sp. PCC 7120 and Nostoc sp. M131, with or without hsdRMS. Strain FD, lacking plasmid D, and an A. variabilis hsdRM deletion mutant were much better hosts for conjugation of a non-replicative, integrative plasmid than wild-type A. variabilis. The conjugative nonreplicating plasmid formed circular molecules in strain FD and in the A. variabilis hsdRM deletion mutant, allowing for a high percentage of single-recombinant exconjugants. In contrast, the relatively few exconjugants in A. variabilis were all double recombinants, suggesting that the restriction-modification system encoded by the genes on plasmid D resulted in only linear molecules that recombined by double crossovers. Expression of the hsdRMS genes in strains Nostoc sp. PCC 7120 and Nostoc sp. M131 drastically reduced conjugation frequency and recombination of a non-replicative plasmid. Together, these data indicate that HsdRMS acts as a defense against the successful transfer of foreign DNA into these cyanobacteria, likely by double-stranded DNA cleavage. Many cyanobacterial strains that are not amenable to gene transfer might have similar restriction-modification systems that lead to cleavage and subsequent degradation of foreign DNA. Although such systems inhibit gene transfer, they may yield a high proportion, although a low number, of gene replacement recombinants.

microbiology↗

bioRxiv2025

Characterization of nitrogenase genes nifP and nifVZT1 in the cyanobacterium Anabaena (Trichormus) variabilis ATCC 29413

Anabaena (Trichormus) variabilis ATCC 29413 is a heterocyst-forming cyanobacterium with two Mo-nitrogenases encoded by the large nif1 and nif2 gene clusters. The nif1-encoded nitrogenase is expressed in heterocysts under oxic growth conditions, whereas the nif2-encoded nitrogenase is expressed only in anoxic vegetative cells. The nifP and nifVZT1 genes are contiguous and distinct from the major nif1 and nif2 clusters, suggesting that nifP and nifVZT1 may be cotranscribed. However, we identified primary transcription start sites for nifP and nifVZT1, indicating that each has its own promoter. The promoter region of nifP shares multiple conserved motifs with the nifB1 and nifB2 promoters, which are regulated by homologous transcriptional activators CnfR1 and CnfR2, respectively. These conserved motifs are not present in the promoter region of nifVZT1. Using nifP promoter fragments that included or excluded the conserved motifs fused to reporter gene lacZ, we determined which motifs contributed to nifP expression; however, these identified motifs differed from the conserved motifs shown previously to regulate nifB1 or nifB2 transcription. Further, the regulation of transcription requires a region inside the coding region of nifP. Mutant strains lacking nifP or nifVZT1 grew well in the absence of fixed nitrogen and showed no reduction in nitrogenase activity compared to the wild-type strain grown under oxic or anoxic conditions. Further, nifP was expressed constitutively in vegetative cells grown -N or +N, but its expression increased greatly in heterocysts by CnfR1 activation of the nifP promoter. In contrast, nifVZT1 was expressed only in heterocysts but, unlike most other nif1 genes, was not under the control of CnfR1.

microbiology↗