Vital roles of the surface layer of the Archaeon Sulfolobus solfataricus: Strong and distinct phenotypes of slaB silenced cultures

Isabelle Anna Zink, Kevin Pfeifer, Erika Wimmer, Christa Schleper

The surface layer (S-layer) is a highly glycosylated proteinaceous lattice which forms the rigid outermost cell envelope of many Archaea and Bacteria. Although it has been implicated to play essential roles in maintaining cell shape and -integrity, protection from environmental stresses and biomineralization, determination of its physiological impact on the living cell has so far been outstanding. The S-layer of the hyperthermophilic Archaeon Sulfolobus solfataricus consists of two proteins: the extracellular SlaA and the transmembrane protein SlaB. Via a novel silencing technique developed in our lab which employs an endogenous RNA-targeting CRISPR type III complex and an artificial mini-CRISPR expressed from a plasmid, we successfully knocked down the slaB gene of S. solfataricus to different low expression levels. Silenced cultures showed strong growth retardation, increase of cell size and distinct surface variations observable by transmission –and scanning electron microscopy and flow cytometry. Notably, specific phenotypes intensified with the level of silencing and recovered upon loss or mutation of the targeting mini-CRISPR or upon complementation via a SlaB of a different Sulfolobusstrain. Moreover, infection assays gave new insights into the involvement of the S-layer in the virus –host interplay. Our work describes for the first time the phenotype of a slaB silenced culture and indicates a vital role of the S-layer in cell size, growth, cell division and integrity and virus infection.

Department für Ökogenomik und Systembiologie
ÖFOS 2012
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