Archaeal transcriptional regulation of the prokaryotic KdpFABC complex mediating K+ uptake in H. salinarum

Abstract

The genome of the halophilic archaeon Halobacterium salinarum encodes the high-affinity ATP-dependent K(+) uptake system Kdp. Previous studies have shown that the genes coding for the KdpFABC complex are arranged in a kdpFABCQ gene cluster together with an additional gene kdpQ. In bacteria, expression of the kdpFABC genes is generally regulated by the dedicated sensor kinase/response regulator pair KdpD/KdpE, which are absent in H. salinarum. Surprisingly, H. salinarum expresses the kdp genes in a manner which is strikingly similar to Escherichia coli. In this study, we show that the halobacterial kdpFABCQ genes constitute an operon and that kdpFABCQ expression is subject to a complex regulatory mechanism involving a negative transcriptional regulator and is further modulated via a so far unknown mechanism. We describe how the regulation of kdp gene expression is facilitated in H. salinarum in contrast to its bacterial counterparts. Whereas the Kdp system fulfils the same core function as an ATP-driven K(+) uptake system in both archaea and bacteria, the different mechanisms involved in the regulation of gene expression appear to have evolved separately, possibly reflecting a different physiological role of ATP-driven K(+) uptake in halophilic archaea.

Construction and Characterization of a Gradually Inducible Expression Vector for Halobacterium salinarum, Based on the kdp Promoter

“…In our previous studies, we showed that expression of the kdpFABCQ operon, which encodes the K ϩ uptake system KdpFABC in H. salinarum, was strictly dependent on the K ϩ concentration in the growth medium (11,18). At extracellular K ϩ concentrations below 20 mM, the kdpFABCQ operon is moderately expressed, whereas expression significantly increased with concentrations below 250 M, with the highest expression level present at a 20 M concentration of external K ϩ .…”

“…At this concentration, the cells ceased to grow due to the affinity limit of the KdpFABC complex for K ϩ . More-detailed studies of the kdp promoter (Pkdp) revealed an operator sequence upstream of the basal transcription elements as well as KdpQ acting as a coregulator stimulating expression under inducing conditions (11,18). Although the particular interplay of these regulatory components is still under investigation, further regulatory components within the kdpFABCQ coding region are clearly absent (11).…”

“…More-detailed studies of the kdp promoter (Pkdp) revealed an operator sequence upstream of the basal transcription elements as well as KdpQ acting as a coregulator stimulating expression under inducing conditions (11,18). Although the particular interplay of these regulatory components is still under investigation, further regulatory components within the kdpFABCQ coding region are clearly absent (11). Plasmid-encoded kdp operon expression from Pkdp resulted in an induction ratio of ϳ50 for noninduced activity versus induced activity (11).…”

“…Although the particular interplay of these regulatory components is still under investigation, further regulatory components within the kdpFABCQ coding region are clearly absent (11). Plasmid-encoded kdp operon expression from Pkdp resulted in an induction ratio of ϳ50 for noninduced activity versus induced activity (11). These findings readily render Pkdp a promising candidate in the establishment of a system in which halobacterial gene expression can be controlled by the simple manipulation of growth conditions (i.e., KCl concentrations).…”

“…These findings readily render Pkdp a promising candidate in the establishment of a system in which halobacterial gene expression can be controlled by the simple manipulation of growth conditions (i.e., KCl concentrations). Since the transcriptional regulatory elements of Pkdp have already been studied in detail (11), this allows a deliberate construction of a versatile expression system. This work deals with the construction and characterization of a replicating vector containing the promoter of the kdpFABCQ operon, thus enabling the controlled inducible expression of target genes in H. salinarum.…”