Biology subjects

Albacar, M.

Publications and source records attributed to Albacar, M..

2 recordsLinked to original sources

bioRxiv2026

Komagataella phaffii encodes two functional Pho4 transcription factors

The transcription factor Pho4 is crucial for the response to phosphate starvation in many fungi, and it has been linked to tolerance to alkalinization of the medium and to pathogenicity. It is widely accepted that it is encoded by a single gene. However, the industrially relevant yeast Komagataella phaffii might contain two Pho4-encoding genes (PAS_chr1-1_0265 and PAS_chr2-1_0177, designated here PHO4(A) and PHO4(B), respectively), which have never been functionally characterized. The phenotypic analysis of single and double mutants suggests that Pho4(B) plays a major role in the adaptation to Pi scarcity. While single mutants exhibited limited and non-overlapping phenotypic defects, the pho4(A) pho4(B) strain was sensitive to multiple types of stress, including phosphate starvation and alkaline pH. Transcriptomic analysis confirms that Pho4(B) is crucial for the transcriptional response to phosphate starvation, including induction of typical gene markers (PHO5, PHO89, VTC1, etc.). However, by using a GFP reporter we found that PHO4(A) also participates in the induction of PHO89 under high pH stress. Expression of both PHO4(A) and PHO4(B) in S. cerevisiae complemented the pho4 mutation under phosphate limitation by restoring growth, expression of the Pho84 transporter and secreted phosphatase activity. These results indicate that both transcription factors display partially overlapping functions, responding differently to diverse stimuli, and that together they constitute a key component in the adaptation to a variety of stresses. Therefore, K. phaffii is an exceptional example among fungi that encodes two Pho4 functional transcription factors.

molecular biology↗

bioRxiv2023

Harnessing alkaline-pH regulatable promoters for efficient methanol-free expression of enzymes of industrial interest in Komagataella phaffii

BackgroundThe yeast Komagataella phaffii has become a very popular host for heterologous protein expression, very often based on the use of the AOX1 promoter, which becomes activated when cells are grown with methanol as a carbon source. However, the use of methanol in industrial settings is not devoid of problems, and therefore, the search for alternative expression methods has become a priority in the last few years. ResultsWe recently reported that moderate alkalinization of the medium triggers a fast and wide transcriptional response in K. phaffii. Here, we present the utilization of three alkaline pH-responsive promoters (pTSA1, pHSP12 and pPHO89) to drive the expression of a secreted phytase enzyme by simply shifting the pH of the medium to 8.0. These promoters offer a wide range of strengths, and the production of phytase could be modulated by adjusting the pH to specific values. The TSA1 and PHO89 promoters offered exquisite regulation, with virtually no enzyme production at acidic pH, while limitation of Pi in the medium further potentiated alkaline pH-driven phytase expression from the PHO89 promoter. An evolved strain based on this promoter was able to produce twice as much phytase as the reference pAOX1-based strain. Functional mapping of the TSA1 and HSP12 promoters suggests that both contain at least two alkaline pH-sensitive regulatory regions. ConclusionsOur work shows that the use of alkaline pH-regulatable promoters could be a useful alternative to methanol-based expression systems, offering advantages in terms of simplicity, safety and economy.

bioengineering↗