Functional characterization of the archaeal-type LonB protease

CERLETTI, M.; MARTINEZ, M.J.; SASTRE, D. E.; GIMÉNEZ M.I.; DE CASTRO RE

Mar del Plata

Congreso; VIII Congreso Argentino de Microbiología General; 2012

Archaea are the third domain of life and their biology has been investigated to a limited extent. Many of the representative members are extremophiles, thus, these unusual microbes are a valuable resource for basic and applied science. Protein turnover is central to cell physiology, however, little is known about protein degradation in archaeal cells. Energy-dependent proteolysis is fundamental for disposal of defective proteins as well as for the regulation of many cellular functions. In Archaea two ATP-dependent proteases have been identified: the soluble proteasome-PAN system and the Lon protease. The Lon protease family is divided into two subfamilies, LonA and LonB. The LonA subfamily consists mainly of bacterial and eukaryotic cytosolic enzymes and accounts for 80% of the presently known Lon proteases while LonB are membrane-bound and predominate in Archaea. The archaeal-type LonB has been studied to a limited extent and the role of this enzyme in archaeal physiology has not been addressed. This study explores the function of the LonB protease in the model haloarchaeon H. volcanii, which flourishes in hypersaline (>2 M NaCl) environments. Lon expression was analyzed at the transcript and protein levels under different conditions. Although relatively constant amounts of Lon protein were detected throughout growth and under the various conditions tested,Lon mRNA levels changed in cells transferred from rich to minimal medium and upon down or up shift in salinity.To further explore about the role of this protease, a deletion mutant (∆HvLon) was constructed in H. volcanii by means of the pop-in-pop-out method. This strain was only obtained when the cells were first complemented with a plasmid expressing the Lon protease in trans, indicating that LonB is an essential protease in H. volcanii and probably in Archaea. Construction of conditional mutants able to produce suboptimal amounts of Lon are in progress to characterize their phenotype. This study provides the first insight into the relevance of the Lon protease in archaeal cells.