The essential role of nitrogen limitation in expression of xplA and degradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in Gordonia sp. strain KTR9.

Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) is a widely used explosive and a major soil and groundwater contaminant. Organisms such as Gordonia sp. KTR9, capable of degrading RDX and using it as an N source, may prove useful for bioremediation of contaminated sites. XplA is a cytochrome P450 monooxygenase responsible for RDX degradation. Expression of xplA in KTR9 was not induced by RDX but was strongly induced (50-fold) during N-limited growth. When glnR, encoding a regulatory protein affecting N assimilation in diverse Actinobacteria, was deleted from KTR9, the bacterium lost the ability to use nitrate, nitrite, and RDX as N sources. Deletion of glnR also abolished the inhibition of xplA expression by nitrite. Our results confirm the essential role of GlnR in regulating assimilation of nitrite, but there was no evidence for a direct role of GlnR in regulating XplA expression. Rather, the general availability of nitrogen repressed XplA expression. We conclude that the inability of the glnR mutant to use RDX as an N source was due to its inability to assimilate nitrite, an intermediate in the assimilation of nitrogen from RDX. Regulation of XplA does not seem adaptive for KTR9, but it is important for RDX bioremediation with KTR9 or similar bacteria.

Regulation of the KstR2 regulon of Mycobacterium tuberculosis by a cholesterol catabolite.

Cholesterol catabolism is widespread in actinobacteria and is critical for Mycobacterium tuberculosis (Mtb) virulence. Catabolism of steroid nucleus rings C and D is poorly understood: it is initiated by the CoA thioesterification of 3aα-H-4α(3'-propanoate)-7aß-methylhexahydro-1,5-indanedione (HIP) by FadD3, whose gene is part of the KstR2 regulon. In Mtb, genes of this regulon were upregulated up to 30- and 22-fold during growth on cholesterol and HIP, respectively, versus another minimal medium. In contrast, genes involved in degrading the cholesterol side-chain and nucleus rings A and B were only upregulated during growth on cholesterol. Similar results were obtained in Rhodococcus jostii RHA1. Moreover, the regulon was not upregulated in a ΔfadD3 mutant unable to produce HIP-CoA. In electrophoretic mobility shift assays, HIP-CoA relieved the binding of KstR2(Mtb) to each of three KstR2 boxes: CoASH, HIP and a related CoA thioester did not. Inspection of the structure of KstR2(RHA1) revealed no obvious HIP-CoA binding pocket. The results establish that Mtb can catabolize the entire cholesterol molecule and that HIP-CoA is an effector of KstR2. They further indicate that KstR2 specifically represses the expression of the HIP degradation genes in actinobacteria, which encode a lower pathway involved in the catabolism of multiple steroids.

Genomic and transcriptomic studies of an RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine)-degrading actinobacterium.

Whole-genome sequencing, transcriptomic analyses, and metabolic reconstruction were used to investigate Gordonia sp. strain KTR9's ability to catabolize a range of compounds, including explosives and steroids. Aspects of this mycolic acid-containing actinobacterium's catabolic potential were experimentally verified and compared with those of rhodococci and mycobacteria.

Photoprotection in the diatom Thalassiosira pseudonana: role of LI818-like proteins in response to high light stress.

As an important component of marine phytoplankton, diatoms must be able to cope with large changes in illumination on a daily basis. They have an active xanthophyll cycle and non-photochemical quenching (NPQ), but no homolog has been detected for the gene encoding the PsbS protein required for NPQ in plants. However, diatoms do have a branch of the light-harvesting complex superfamily, the Lhcx clade, which is most closely related to the LI818 (LhcSR) genes of the green alga Chlamydomonas, known to be upregulated in response to a variety of stresses. When cultures of the diatom T. pseudonana grown under low light (40 micromol photons m(-2) s(-1)) were exposed to high light stress (HL, 700 micromol photons m(-2) s(-1)), transcripts of three of these genes (Lhcx1, Lhcx4, Lhcx6) were transiently accumulated. The amount of Lhcx6 protein was low under low light, but increased continuously during 10h of HL exposure, then slowly dropped to background levels in the dark. However, HL had little effect on the Lhcx1 protein, which was present under low light and only doubled after HL exposure. Diatoxanthin levels increased throughout the HL period with no change in diadinoxanthin. The fraction of NPQ attributable to photoinhibitory quenching (qI) also increased throughout the HL exposure. Taken together, the Lhcx6 protein could be associated with diatoxanthin binding and play a direct role in excess energy dissipation via sustained quenching during acclimation to prolonged HL stress, while the Lhcx1 protein may play a more structural role in thylakoid membrane organization under all conditions.