The Saccharomyces cerevisiae Ncw2 protein works on the chitin/ß-glucan organisation of the cell wall.

The NCW2 gene was recently described as encoding a GPI-bounded protein that assists in the re-modelling of the Saccharomyces cerevisiae cell wall (CW) and in the repair of damage caused by the polyhexamethylene biguanide (PHMB) polymer to the cell wall. Its absence produces a re-organization of the CW structure that result in resistance to lysis by glucanase. Hence, the present study aimed to extend the analysis of the Ncw2 protein (Ncw2p) to determine its physiological role in the yeast cell surface. The results showed that Ncw2p is transported to the cell surface upon O-mannosylation mediated by the Pmt1p-Pmt2p enzyme complex. It co-localises with the yeast bud scars, a region in cell surface formed by chitin deposition. Once there, Ncw2p enables correct chitin/ß-glucan structuring during the exponential growth. The increase in molecular mass by hyper-mannosylation coincides with the increasing in chitin deposition, and leads to glucanase resistance. Treatment of the yeast cells with PHMB produced the same biological effects observed for the passage from exponential to stationary growth phase. This might be a possible mechanism of yeast protection against cationic biocides. In conclusion, we propose that Ncw2p takes part in the mechanism involved in the control of cell surface rigidity by aiding on the linkage between chitin and glucan layers in the modelling of the cell wall during cell growth.

Lawsone, a 2-hydroxy-1,4-naphthoquinone from Lawsonia inermis (henna), produces mitochondrial dysfunctions and triggers mitophagy in Saccharomyces cerevisiae.

Lawsone is a natural naphthoquinone present in the henna leaf extract with several cytotoxic activities and used as precursor for synthesis of various pharmaceutical compounds. Its biological activities are thought to be the result of oxidative stress generated, although the hydroxy group at position C-2 in its structure tends to reduce its electrophilic potential. In view of lack of knowledge on its activity, the present work aimed to elucidate the biological effect of lawsone using the yeast Saccharomyces cerevisiae. In the model strain BY4741 it was defined 229 mmol/L as the minimal inhibitory concentration (MIC). Using 172 mmol/L as sub-MIC value it was observed that yap1 deletion mutant was sensitive to lawsone independent the presence of oxygen. Lawsone affected yeast growth in glycerol, indicating interference in the respiratory metabolism. Intracellular content of thiol groups did not indicate intensive oxidative stress and the presence of the anti-oxidant N-acetylcysteine (NAC) exacerbated lawsone toxicity. By analysing the sensitivity of atg mutant strains and the localization of GFP-Atg8 fusion protein, it was concluded that lawsone primarily produces mitochondrial malfunctioning, leading to indirect oxidative stress. It triggers the autophagic response that ultimately induces mitophagy.

The effects of the Ncw2 protein of Saccharomyces cerevisiae on the positioning of chitin in response to cell wall damage.

The recently described NCW2 gene encodes a protein that is assumed to be located in the cell wall (CW). This protein was proposed to participate in the repair of CW damages induced by polyhexamethylene biguanide (PHMB). However, much of the information on the biological function(s) of Ncw2p still remains unclear. In view of this, this study seeks to extend the analysis of this gene in light of the way its protein functions in the Cell Wall Integrity (CWI) mechanism. Deletion of the NCW2 gene led to constitutive overexpression of some key CWI genes and increased chitin deposition in the walls of cells exposed to PHMB. This means the lack of Ncw2p might activate a compensatory mechanism that upregulates glucan CWI genes for cell protection by stiffening the CW. This condition seems to alleviate the response through the HOG pathway and makes cells sensitive to osmotic stress. However, Ncw2p may not have been directly involved in tolerance to osmotic stress itself. The results obtained definitely place the NCW2 gene in the list of CWI genes of S. cerevisiae and indicate that its protein has an auxiliary function in the maintenance of the glucan/chitin balance and ensuring the correct structure of the yeast cell wall.

Complete nucleotide sequences of two blaKPC-2-bearing IncN Plasmids isolated from sequence type 442 Klebsiella pneumoniae clinical strains four years apart.

We sequenced the oldest blaKPC-2-bearing plasmid isolated in Brazil and another plasmid also carried by a Klebsiella pneumoniae strain of sequence type 442 (ST442), isolated 52 months later. Both plasmids present an IncN backbone and few acquired regions. Because the 2005 plasmid presented deletions and a truncated gene within Tn4401b compared to the 2009 plasmid, we can thus infer that IncN blaKPC-2-bearing plasmids pFCF1305 and pFCF3SP had a common ancestor circulating in Brazil prior to May 2005.