Unravelling archaeal signal transduction networks
Final Report Abstract
Crenarchaea employ the reversible phosphorylation of serine, threonines and tyrosines for signaling purposes in different cellular networks. One of the best studied pathways is the regulation of the archaellum regulon which expresses the motility machinery of archaea. Numerous regulators have been found to play a role in the regulation of the archaellum regulon in Sulfolobus acidocaldarius, to which a set of protein kinases and the serine/threonine phosphatase PP2A belong. During the funding period we have uncovered more regulators that are involved in the modulation of expression of the archaellum regulon in S. acidocaldarius. Phosphorylation and dephosphorylation of regulators in this process seems to play an important part for their function as after starvation four regulators formed a very stable complex with the serine/threonine protein phosphatase PP2A. These were ArnA/B, the already known inhibitors of archaellum gene expression; UspA, a universal stress protein and the SaGPN GTPase, which turned out to be a positive regulator of archaellum gene expression. We characterized the interaction of ArnA/B in more detail and found that it is phosphorylation dependent. Dephosphorylation by PP2A destabilized the complex. Possibly thereby leading to a relief of the inhibition of the archaellin promoter and allowing for transcription of the arlB gene. This might coincide with the presence of the SaGPN in the complex as this protein seems to be important for the induction of arlB transcription. This might be due to binding to the PP2A, which in turn then is able to interact with ArnA/B. We are currently trying to study these effects in vitro with purified enzymes. The function of UspA in this process remains unclear. We now ask the question whether the PP2A in S. acidocaldarius is like the ones in eukarya which act as interaction hubs for a variety of proteins involved in cellular signaling pathways. We showed that the protein is expressed at similar levels at all growth conditions. Therefore, its activity is most likely modulated by different interaction partners at different conditions. We are currently performing pull-down experiments at different growth conditions to analyze whether the interacting proteins differ or whether the here described complex is always associated with the PP2A. The archaellum regulatory network has become even more complex with this study and we will need to analyze the interaction of the different factors in more detail to get a better understanding of the different levels of regulation.
Publications
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(2019) Structure and interactions of the archaeal motility repression module ArnA-ArnB that modulates archaellum gene expression in Sulfolobus acidocaldarius. J Biol Chem 294:7460-7471
Hoffmann L, Anders K, Bischof LF, Ye X, Reimann J, Khadouma S, Pham TK, van der Does C, Wright PC, Essen LO, Albers SV
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(2020) SaUspA, the Universal Stress Protein of Sulfolobus acidocaldarius stimulates the activity of the PP2A Phosphatase and is involved in growth at high salinity. Front Microbiol 11:598821
Ye X, van der Does C, Albers SV
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(2020) The Phosphatase PP2A Interacts With ArnA and ArnB to Regulate the Oligomeric State and the Stability of the ArnA/B Complex. Front Microbiol. 11:1849
Ye X, Vogt MS, van der Does C, Bildl W, Schulte U, Essen LO, Albers SV
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(2022). Methods for markerless gene deletion and plasmid-based expression in Sulfolobus acidocaldarius. Methods Mol Biol 2522:135-144
Ye X, Recalde A, Albers SV, van Wolferen
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(2022). Methods to analyze motility in eury- and crenarchaea. Methods Mol Biol 2522:373-385
Patro M, van Wolferen M, Ye X, Albers SV and Quax TEF
