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.

Many extremely halophilic archaea belonging to the Halobacteriales have remarkable longevity. They are even known to persist for millions of years within fluid inclusions of salt crystals. However, the key systems responsible for this remarkable ability and the underlying physiological mechanisms have not yet been deciphered. This study revealed that the ATP-dependent K(+) uptake system KdpFABC of Halobacterium salinarum is essential for survival under desiccation and salt crystal inclusion and, thus, can be identified as at least one of these systems in this organism. The presence of the kdp genes promoted survival of H. salinarum entombed in halite, compared with cells in which these genes were deleted. Expression of the kdp operon was found to be induced already under desiccating conditions without halite entombment. The morphology of cells included in halite resembled that of cells grown under potassium limitation. Therefore, a steady potassium supply, even under unfavourable energetic conditions, plays a key role in long-term survival and desiccation tolerance.

Gradually inducible expression vectors which are governed by variations of growth conditions are powerful tools for gene expression of conditionally lethal mutants. Furthermore, controlled expression allows monitoring of overproduction of proteins at various stages in their expressing hosts. For Halobacterium salinarum, which is often used as a paradigm for halophilic archaea, such an inducible expression system is not available to date. Here we show that the kdp promoter (Pkdp), which facilitates gene expression upon K ؉ limitation, can be used to establish such a system for molecular applications. Pkdp features a rather high expression rate, with an approximately 50-fold increase that can be easily varied by K ؉ concentrations in the growth medium. Besides the construction of an expression vector, our work describes the characterization of expression patterns and, thus, offers a gradually inducible expression system to the scientific community.

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