CaGdt1 plays a compensatory role for the calcium pump CaPmr1 in the regulation of calcium signaling and cell wall integrity signaling in Candida albicans

Abstract

BackgroundSaccharomyces cerevisiae ScGdt1 and mammalian TMEM165 are two members of the UPF0016 membrane protein family that is likely to form a new group of Ca2+/H+ antiporter and/or a Mn2+ transporter in the Golgi apparatus. We have previously shown that Candida albicans CaGDT1 is a functional ortholog of ScGDT1 in the response of S. cerevisiae to calcium stress. However, how CaGdt1 together with the Golgi calcium pump CaPmr1 regulate calcium homeostasis and cell wall integrity in this fungal pathogen remains unknown.MethodsChemical sensitivity was tested by dilution assay. Cell survival was examined by measuring colony-forming units and staining with Annexin V-FITC and propidium iodide. Calcium signaling was examined by expression of downstream target gene CaUTR2, while cell wall integrity signaling was revealed by detection of phosphorylated Mkc1 and Cek1. Subcellular localization of CaGdt1 was examined through direct and indirect immunofluorescent approaches. Transcriptomic analysis was carried out with RNA sequencing.ResultsThis study shows that Candida albicans CaGDT1 is also a functional ortholog of ScGDT1 in the response of S. cerevisiae to cell wall stress. CaGdt1 is localized in the Golgi apparatus but at distinct sites from CaPmr1 in C. albicans. Loss of CaGDT1 increases the sensitivity of cell lacking CaPMR1 to cell wall and ER stresses. Deletion of CaGDT1 and/or CaPMR1 increases calcium uptake and activates the calcium/calcineurin signaling. Transcriptomic profiling reveals that core functions shared by CaGdt1 and CaPmr1 are involved in the regulation of cellular transport of metal ions and amino acids. However, CaGdt1 has distinct functions from CaPmr1. Chitin synthase gene CHS2 is up regulated in all three mutants, while CHS3 is only up regulated in the pmr1/pmr1 and the gdt1/gdt1 pmr1/pmr1 mutants. Five genes (DIE2, STT3, OST3, PMT1 and PMT4) of glycosylation pathway and one gene (SWI4) of the cell wall integrity (CWI) pathway are upregulated due to deletion of CaGDT1 and/or CaPMR1. Consistently, deletion of either CaPMR1 or CaGDT1 activates the CaCek1-mediated CWI signaling in a cell wall stress-independent fashion. Calcineurin function is required for the integrity of the cell wall and vacuolar compartments of cells lacking both GDT1 and CaPMR1.ConclusionsCaPmr1 is the major player in the regulation of calcium homeostasis and cell wall stress, while CaGdt1 plays a compensatory role for CaPmr1 in the Golgi compartment in C. albicans.Electronic supplementary materialThe online version of this article (10.1186/s12964-018-0246-x) contains supplementary material, which is available to authorized users.

The protein kinase Cmk2 negatively regulates the calcium/calcineurin signalling pathway and expression of calcium pump genes PMR1 and PMC1 in budding yeast

“…The PMC1 - lac Z reporter and PMR1 - lac Z reporter were described in our previous study [28]. The β-galactosidase activity was determined using the substrate ONPG as described [29–31]. Data are mean ± SD from six independent experiments.…”

RNA sequencing reveals an additional Crz1-binding motif in promoters of its target genes in the human fungal pathogen Candida albicans

“…To identify genes regulated by CaCrz1, the wild type SN148 and its isogenic CRISPR mutant for CaCRZ1 were grown to log-phase at 30°C before they were treated with 0.2 M CaCl 2 for 2 h. Total RNA samples were extracted Qiagen RNeasy minikit protocol, and RNA integrity was evaluated using an Agilent 2100 Bioanalyzer (Agilent Technologies, USA) as described [45]. RNA-seq libraries were constructed using Illumina's miSEQ RNA Sample Preparation Kit (Illumina Inc., USA).…”

From the Uncharacterized Protein Family 0016 to the GDT1 family: Molecular insights into a newly-characterized family of cation secondary transporters

“…In the yeast C. albicans , the comparison of the double deletion strain to each of the single deletions indicates that CaGdt1p and CaPmr1p also interact at the genetic level. Indeed, further deletion of CaGDT1 in a strain deleted for CaPMR1 increases (i) its sensitivity towards cell wall and ER stresses, (ii) its ability to accumulate radioactive 45 Ca 2+ , and (iii) its hypersensitivity to inhibitors of the Ca 2+ -mediated calcineurin signaling pathway [ 24 ]. Transcriptomic analyses further revealed that CaGdt1p is involved in the regulation of cellular transport of metal ions and amino acids [ 24 ].…”

“…Indeed, further deletion of CaGDT1 in a strain deleted for CaPMR1 increases (i) its sensitivity towards cell wall and ER stresses, (ii) its ability to accumulate radioactive 45 Ca 2+ , and (iii) its hypersensitivity to inhibitors of the Ca 2+ -mediated calcineurin signaling pathway [ 24 ]. Transcriptomic analyses further revealed that CaGdt1p is involved in the regulation of cellular transport of metal ions and amino acids [ 24 ]. In this pathogen, CaGdt1p also interacts at the genetic level with the high-affinity plasma membrane Ca 2+ channel CaCch1p/CaMid1p, again strengthening the role of CaGdt1p in Ca 2+ homeostasis.…”