Genetic Interactions Between Regulators of Chlamydomonas Phosphorus and Sulfur Deprivation Responses

Abstract

The Chlamydomonas reinhardtii PSR1 gene is required for proper acclimation of the cells to phosphorus (P) deficiency. P-starved psr1 mutants show signs of secondary sulfur (S) starvation, exemplified by the synthesis of extracellular arylsulfatase and the accumulation of transcripts encoding proteins involved in S scavenging and assimilation. Epistasis analysis reveals that induction of the S-starvation responses in P-limited psr1 cells requires the regulatory protein kinase SNRK2.1, but bypasses the membrane-targeted activator, SAC1. The inhibitory kinase SNRK2.2 is necessary for repression of S-starvation responses during both nutrient-replete growth and P limitation; arylsulfatase activity and S deficiency-responsive genes are partially induced in the P-deficient snrk2.2 mutants and become fully activated in the P-deficient psr1snrk2.2 double mutant. During P starvation, the sac1snrk2.2 double mutants or the psr1sac1snrk2.2 triple mutants exhibit reduced arylsulfatase activity compared to snrk2.2 or psr1snrk2.2, respectively, but the sac1 mutation has little effect on the abundance of S deficiency-responsive transcripts in these strains, suggesting a post-transcriptional role for SAC1 in elicitation of S-starvation responses. Interestingly, P-starved psr1snrk2.2 cells bleach and die more rapidly than wild-type or psr1 strains, suggesting that activation of S-starvation responses during P deprivation is deleterious to the cell. From these results we infer that (i) P-deficient growth causes some internal S limitation, but the S-deficiency responses are normally inhibited during acclimation to P deprivation; (ii) the S-deficiency responses are not completely suppressed in P-deficient psr1 cells and consequently these cells synthesize some arylsulfatase and exhibit elevated levels of transcripts for S-deprivation genes; and (iii) this increased expression is controlled by regulators that modulate transcription of S-responsive genes during S-deprivation conditions. Overall, the work strongly suggests integration of the different circuits that control nutrient-deprivation responses in Chlamydomonas.T HE elements phosphorus (P) and sulfur (S) are essential macronutrients for sustaining life. P is a structural component of nucleic acids and phospholipids, and is a ubiquitous modifier of carbohydrates and proteins, while S is incorporated into sulfolipids, polysaccharides, proteins, cofactors, and a wide variety of important metabolites including S-adenosyl-methionine, glutathione, and phytochelatins. The preferred forms of P and S that are assimilated by plants and microbes are the orthophosphate ion, PO 4 3À (Pi), and the sulfate ion, SO 4 2À

Identification and Regulation of Plasma Membrane Sulfate Transporters in Chlamydomonas

“…In contrast to wild-type cells, the sac1 mutant only accumulates SLT1, SLT2, and SULTR2 transcripts 24 h after the imposition of S starvation. This is consistent with the model proposed by Moseley et al (2009), in which SAC1 is a sensor that resides on the plasma membrane and acts as a negative regulator of the SNRK2.2 repressor protein kinase when environmental SO 4 22 levels fall below a certain threshold. This causes derepression of the S-responsive genes, including those encoding SO 4 22 transporters.…”

Sulphur responsiveness of the Chlamydomonas reinhardtii LHCBM9 promoter

“…This supported the findings of previous work, where the LHCBM9 gene was not found to be up-regulated upon N deprivation Miller et al 2007;Moseley et al 2009), but contradicted a recent study by Grewe et al (2014), who found that N deprivation induced LHCBM9 protein expression. It is possible that LHCBM9 is regulated by N deprivation at a level other than transcription, especially since N-dependent gene regulation shares regulatory elements with the S-response pathway (Kopriva and Rennenberg 2004;Shin 2011).…”

Tiered Regulation of Sulfur Deprivation Responses in Chlamydomonas reinhardtii and Identification of an Associated Regulatory Factor

“…B and C, RT-qPCR showing the levels of SULTR2, SLT1, SLT2, AOT4, SULTR1, ARS1, ARS2, ECP76, SBDP, LHCBM9, HAP2, and OASTL transcripts in the snrk2.2 single mutant (B) and snrk2.2 ars73a double mutant (C). Davies et al, 1994Davies et al, , 1996Davies et al, , 1999Yildiz et al, 1994Yildiz et al, , 1996Takahashi et al, 2001;Zhang et al, 2004;Chang et al, 2005;González-Ballester et al, 2008Moseley et al, 2009;Pootakham et al, 2010), and one of the first responses of this alga to S limitation was shown to be the activation of genes encoding high-affinity SO 4 22 transporters González-Ballester et al, 2010;Pootakham et al, 2010). Increased expression from high-affinity SO 4 22 transporters has been noted to occur within 30 min of the removal of S from the culture medium ( Fig.…”