Carmen-Lisset Flores: "Una proteína pluriempleada en la levadura no convencional Yarrowia lipolytica"

23/01/2020

Moonlighting proteins are able to perform different unrelated functions and are not theresult of gene fusion.. We have uncovered a protein in the non-conventional yeast Yarrowia lipolytica that shows characteristics of a moonlighting protein. This protein is the N-acetylglucosamine (NAGA) kinase, the first intracellular protein in the metabolism of NAGA. This metabolism implicates the coordinated expression of 4 genes (NAGgenes) encoding respectively a transporter (NGT1), the kinase (NAG5), a deacetylase (NAG2) and a deaminase (NAG1). The final product of the metabolic pathway is fructose-6-P that enters the glycolytic trunk. Growth in NAGA promotes the transcription of these genes while grow in glucose abolishes it. However, disruption of the gene YlNAG5that causes inability to grow in NAGA, allows transcription of the NAGgenes during growth in glucose. This suggests that the YlNag5 protein could have two functions: one as kinase,the other as transcriptional regulator. To test this hypothesis we have expressed several heterologous NAGA kinases or modified versions of the YlNag5 protein in an Ylnag5mutant and examined if they complemented both functions. Expression of these proteins was checked by western blot. NAGA kinase activity was measured and transcription state of the genes was assessed by determining the enzymatic activity of YlNag1. We found that proteins with similar kinase activity behaved differently in their role as transcriptional regulator thus showing that these functions may be separated. A search for proteins that could interact with YlNag5, using a pulldown assay, showed that YlNag5 interacts with a protein similar to Saccharomyces cerevisiae Urp2, a component of the small (40S) ribosomal subunit.An alternative explanation for the behaviour of the Ylnag5mutants could be that the deregulation of the NAGgenes were triggered by NAGA accumulation in the mutant. Chitin hydrolysis, which takes place outside the cell, would be the source of NAGA in this case. We found that in a double mutant Ylnag5 ngt1 that cannot internalize NAGA, the deaminase activity is derepressed. Las levaduras: en la intersección entre la Biología de sistemas y la BiomedicinaEnmemoriadelProfesorJulioRodríguezVillanuevaYeasts: at the cross-roads of Systems biology and BiomedicineIn memory of Professor Julio Rodríguez VillanuevaMadrid, 23 y 24 de enero de 2020 / January, 23 and 24, 2020 Our results support the idea that YlNag5 is a moonlighting protein with a metabolic and a regulatory role in the transcription of the NAGgenes.Acknowledgments.-This work was supported by grant CIVP18A3896 of the Fundación Ramón Areces.

Vídeos
Ciencias de la vida y de la materia las-levaduras-en-la-interseccion-entre-la-biologia-de-sistemas-y-la-biomedicina-en-memoria-del-profesor-julio-rodriguez-villanueva

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