Innokentev Aleksei, Sanchez Ana M, Monetti Mara, Schwer Beate, Shuman Stewart
Molecular Biology Program, Memorial Sloan Kettering Cancer Center, New York, New York, USA.
Gerstner Sloan Kettering Graduate School of Biomedical Sciences, New York, New York, USA.
mBio. 2025 Jan 8;16(1):e0299224. doi: 10.1128/mbio.02992-24. Epub 2024 Dec 11.
The fission yeast regulon genes , , and -encoding a cell surface-associated acid phosphatase (Pho1), a plasma membrane inorganic phosphate transporter (Pho84), and a plasma membrane glycerophosphocholine transporter (Tgp1)-are strongly upregulated in response to acute phosphate starvation, as are the and genes that encode putative 5'-nucleotidase paralogs of the binuclear metallophosphoesterase enzyme superfamily. Via proteomic analysis of the medium harvested from phosphate-replete and phosphate-starved fission yeast, we define a starvation secretome that includes SPBPB2B2.06c (renamed Efn1, for xtracellular ive-prime ucleotidase), SPAC1039.02 (henceforth Efn2), and Pho1 among the most abundant exported proteins elaborated by phosphate-starved cells. We demonstrate and characterize a 5'-nucleotidase activity secreted into the medium of phosphate-starved cells, which is eliminated by simultaneous deletion of and . By singly deleting and , we find that Efn1 contributes the greater share of secreted 5'-nucleotidase activity. Efn1 and Efn2 catalyze the release of inorganic phosphate from all four standard ribonucleoside monophosphates, in order of preference: CMP > UMP > AMP > GMP. Whereas cells can use extracellular CMP as a source of phosphate during phosphate starvation, ∆ ∆ cells cannot. The secretion of 5'-nucleotidase enzymes during phosphate limitation is a newly appreciated facet of fission yeast phosphate homeostasis.
adapts to phosphate starvation by upregulating the expression of a cell surface acid phosphatase that mobilizes inorganic phosphate from the extracellular milieu, as well as transmembrane transporters that take up inorganic phosphate and glycerophosphocholine. This study identifies two paralogous extracellular 5'-nucleotidase enzymes, Efn1 and Efn2, encoded by genes that are highly transcriptionally induced during acute phosphate starvation, as major proteins secreted into the medium by phosphate-starved fission yeast cells. Secreted Efn1 and Efn2 catalyze the release of inorganic phosphate from all ribonucleoside monophosphates, with a preference for CMP. Secretion of Efn1 and Efn2 enables phosphate-starved fission yeast to thrive by using extracellular CMP as a source of inorganic phosphate. The starvation-induced production of extracellular 5'-nucleotidases adds a new layer of pro-adaptive function during phosphate limitation.
裂殖酵母调节子基因、和——分别编码一种细胞表面相关酸性磷酸酶（Pho1）、一种质膜无机磷酸盐转运蛋白（Pho84）和一种质膜甘油磷酸胆碱转运蛋白（Tgp1）——在急性磷酸盐饥饿时会强烈上调表达，编码双核金属磷酸二酯酶超家族假定5'-核苷酸酶旁系同源物的和基因也是如此。通过对从富含磷酸盐和缺乏磷酸盐的裂殖酵母中收获的培养基进行蛋白质组分析，我们确定了一个饥饿分泌组，其中包括SPBPB2B2.06c（重命名为Efn1，即细胞外5'-核苷酸酶）、SPAC1039.02（此后称为Efn2）以及Pho1，它们是缺乏磷酸盐的细胞分泌的最丰富的胞外蛋白。我们证明并表征了一种分泌到缺乏磷酸盐的细胞培养基中的5'-核苷酸酶活性，同时缺失和会消除这种活性。通过单独缺失和，我们发现Efn1对分泌的5'-核苷酸酶活性贡献更大。Efn1和Efn2催化从所有四种标准核糖核苷单磷酸中释放无机磷酸盐，偏好顺序为：CMP > UMP > AMP > GMP。虽然细胞在磷酸盐饥饿期间可以利用细胞外CMP作为磷酸盐来源，但∆∆细胞则不能。在磷酸盐限制期间5'-核苷酸酶酶的分泌是裂殖酵母磷酸盐稳态一个新认识的方面。
通过上调一种从细胞外环境中动员无机磷酸盐的细胞表面酸性磷酸酶以及摄取无机磷酸盐和甘油磷酸胆碱的跨膜转运蛋白的表达来适应磷酸盐饥饿。本研究确定了两种同源的细胞外5'-核苷酸酶Efn1和Efn2，它们由在急性磷酸盐饥饿期间高度转录诱导的基因编码，是缺乏磷酸盐的裂殖酵母细胞分泌到培养基中的主要蛋白质。分泌的Efn1和Efn2催化从所有核糖核苷单磷酸中释放无机磷酸盐，偏好CMP。Efn1和Efn2的分泌使缺乏磷酸盐的裂殖酵母能够通过利用细胞外CMP作为无机磷酸盐来源而茁壮成长。饥饿诱导的细胞外5'-核苷酸酶的产生在磷酸盐限制期间增加了一层新的适应性功能。
