Membrane Interactome of a Recombinant Fragment of Human Surfactant Protein D Reveals GRP78 as a Novel Binding Partner in PC3, a Metastatic Prostate Cancer Cell Line

TY - JOUR

T1 - Membrane Interactome of a Recombinant Fragment of Human Surfactant Protein D Reveals GRP78 as a Novel Binding Partner in PC3, a Metastatic Prostate Cancer Cell Line

AU - Thakur, Gargi

AU - Sathe, Gajanan

AU - Kundu, Indra

AU - Biswas, Barnali

AU - Gautam, Poonam

AU - Alkahtani, Saad

AU - Idicula-Thomas, Susan

AU - Sirdeshmukh, Ravi

AU - Kishore, Uday

AU - Madan, Taruna

N1 - Funding Information: We thank ICMR and Dr. Smita Mahale, Director, ICMR-National Institute for Research in Reproductive Health (ICMR-NIRRH), Mumbai, for intramural funding (Accession number 921). We thank Mr. Vaibhav Shinde for his support to enhance resolution of the figures. SA was funded by Researchers Supporting Project (RSP-2020/26) King Saud University, Riyadh. Publisher Copyright: © Copyright © 2021 Thakur, Sathe, Kundu, Biswas, Gautam, Alkahtani, Idicula-Thomas, Sirdeshmukh, Kishore and Madan.

PY - 2021/1/19

Y1 - 2021/1/19

N2 - Surfactant protein-D (SP-D), a member of the collectin family has been shown to induce apoptosis in cancer cells. SP-D is composed of an N-terminal collagen-like domain and a calcium-dependent carbohydrate recognition domain (CRD). Recently, we reported that a recombinant fragment of human SP-D (rfhSP-D), composed of homotrimeric CRD region, induced intrinsic apoptotic pathway in prostate cancer cells. Here, we analyzed the membrane interactome of rfhSP-D in an androgen-independent prostate cancer cell line, PC3, by high resolution mass spectrometry and identified 347 proteins. Computational analysis of PPI network of this interactome in the context of prostate cancer metastasis and apoptosis revealed Glucose Regulated Protein of 78 kDa (GRP78) as an important binding partner of rfhSP-D. Docking studies suggested that rfhSP-D (CRD) bound to the substrate-binding domain of glycosylated GRP78. This was further supported by the observations that human recombinant GRP78 interfered with the binding of rfhSP-D to anti-SP-D polyclonal antibodies; GRP78 also significantly inhibited the binding of recombinant full-length human SP-D with a monoclonal antibody specific to the CRD in a dose-dependent manner. We conclude that the interaction with rfhSP-D is likely to interfere with the pro-survival signaling of GRP78.

AB - Surfactant protein-D (SP-D), a member of the collectin family has been shown to induce apoptosis in cancer cells. SP-D is composed of an N-terminal collagen-like domain and a calcium-dependent carbohydrate recognition domain (CRD). Recently, we reported that a recombinant fragment of human SP-D (rfhSP-D), composed of homotrimeric CRD region, induced intrinsic apoptotic pathway in prostate cancer cells. Here, we analyzed the membrane interactome of rfhSP-D in an androgen-independent prostate cancer cell line, PC3, by high resolution mass spectrometry and identified 347 proteins. Computational analysis of PPI network of this interactome in the context of prostate cancer metastasis and apoptosis revealed Glucose Regulated Protein of 78 kDa (GRP78) as an important binding partner of rfhSP-D. Docking studies suggested that rfhSP-D (CRD) bound to the substrate-binding domain of glycosylated GRP78. This was further supported by the observations that human recombinant GRP78 interfered with the binding of rfhSP-D to anti-SP-D polyclonal antibodies; GRP78 also significantly inhibited the binding of recombinant full-length human SP-D with a monoclonal antibody specific to the CRD in a dose-dependent manner. We conclude that the interaction with rfhSP-D is likely to interfere with the pro-survival signaling of GRP78.

KW - apoptosis

KW - GRP78

KW - innate immunity

KW - interactome analysis

KW - prostate cancer

KW - signaling

KW - surfactant protein D

UR - https://www.scopus.com/pages/publications/85100524756

U2 - 10.3389/fimmu.2020.600660

DO - 10.3389/fimmu.2020.600660

M3 - Article

C2 - 33542717

AN - SCOPUS:85100524756

SN - 1664-3224

VL - 11

JO - Frontiers in Immunology

JF - Frontiers in Immunology

M1 - 600660

ER -