Prostate secretory protein 94 inhibits sterol binding and export by the mammalian CAP protein CRISP2 in a calcium-sensitive manner

Ola El Atab,Aslihan Ekim Kocabey,Oluwatoyin A Asojo,Roger Schneiter

doi:10.1016/j.jbc.2022.101600

Ola El Atab, Aslihan Ekim Kocabey + Show 2 more

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https://doi.org/10.1016/j.jbc.2022.101600

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Abstract

Members of the CAP protein superfamily are present in all kingdoms of life and have been implicated in many different processes, including pathogen defense, immune evasion, sperm maturation, and cancer progression. Most CAP proteins are secreted glycoproteins and share a unique conserved αβα sandwich fold. The precise mode of action of this class of proteins, however, has remained elusive. Saccharomyces cerevisiae has three CAP family members, termed pathogen related in yeast (Pry). We have previously shown that Pry1 and Pry2 export sterols in vivo and that they bind sterols in vitro. This sterol binding and export function of yeast Pry proteins is conserved in the mammalian CRISP proteins and other CAP superfamily members. CRISP3 is an abundant protein of the human seminal plasma and interacts with prostate secretory protein of 94 amino acids (PSP94), another major protein component in the seminal plasma. Here we examine whether the interaction between CRISP proteins and PSP94 affects the sterol binding function of CAP family members. We show that coexpression of PSP94 with CAP proteins in yeast abolished their sterol export function and the interaction between PSP94 and CAP proteins inhibits sterol binding in vitro. In addition, mutations that affect the formation of the PSP94–CRISP2 heteromeric complex restore sterol binding. Of interest, we found the interaction of PSP94 with CRISP2 is sensitive to high calcium concentrations. The observation that PSP94 modulates the sterol binding function of CRISP2 in a calcium-dependent manner has potential implications for the role of PSP94 and CRISP2 in prostate physiology and progression of prostate cancer.

Highlights

CAP proteins form a large superfamily with members found in all kingdoms of life.The superfamily was named after the identification of sequence similarities between its founding members: CRISPs (Cysteine-rich secretory proteins) of the mammalian reproductive tract, Ag5 (Antigen 5) in the venom secretory duct of stinging insects and PR-1 (PathogenesisRelated protein-1) which is induced in plants upon pathogen infection
To test whether expression of Prostate secretory protein 94 (PSP94) would affect the function of Pry1 in sterol export, we expressed PSP94 fused to the N-terminal signal sequence from Pry1 from a constitutive alcohol dehydrogenase promoter (ADH1) to direct the protein into the yeast secretory pathway
To test whether PSP94 affects fatty acid binding by the CAP domain, we examined whether coexpression of PSP94 with CRISP2 affects fatty acid export from mutant cells lacking the two acyl-CoA synthases Faa1 and Faa4, and two of the 3 yeast CAP domain proteins, Pry1 and Pry3

Summary

Introduction

The superfamily was named after the identification of sequence similarities between its founding members: CRISPs (Cysteine-rich secretory proteins) of the mammalian reproductive tract, Ag5 (Antigen 5) in the venom secretory duct of stinging insects and PR-1 (PathogenesisRelated protein-1) which is induced in plants upon pathogen infection. They are referred to as Sperm Coating Proteins (SCP) or TAPS (Tpx-1/Ag5/PR-1/Sc7). Journal Pre-proof three-stranded anti-parallel -sheet flanked by -helices on both sides [1] They are mostly secreted glycoproteins, implicated in many fundamental biological processes including immune defense in mammals and plants, sperm maturation and fertilization, prostate and brain cancer, pathogen virulence and venom toxicity. Even though these proteins are extensively studied, their precise mode of action remains to be defined [2,3,4]

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