Additional file 1 of PGRMC1 phosphorylation affects cell shape, motility, glycolysis, mitochondrial form and function, and tumor growth
Creators: name: Thejer, Bashar M.
name: Partho P. Adhikary
name: Amandeep Kaur
name: Teakel, Sarah L.
name: Oosterum, Ashleigh Van
name: Ishith Seth
name: Pajic, Marina
name: Hannan, Katherine M.
name: Pavy, Megan
name: Perlita Poh
name: Jazayeri, Jalal A.
name: Thiri Zaw
name: Pascovici, Dana
name: Ludescher, Marina
name: Pawlak, Michael
name: Cassano, Juan C.
name: Turnbull, Lynne
name: Jazayeri, Mitra
name: James, Alexander C.
name: Coorey, Craig P.
name: Roberts, Tara L.
name: Kinder, Simon J.
name: Hannan, Ross D.
name: Patrick, Ellis
name: Molloy, Mark P.
name: New, Elizabeth J.
name: Fehm, Tanja N.
name: Neubauer, Hans
name: Goldys, Ewa M.
name: Weston, Leslie A.
name: Cahill, Michael A.
Publisher: name: figshare
Year: 2020
Subjects: subject: Biophysics
subject: Biochemistry
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Physiology
subject: Pharmacology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Developmental Biology
subject: Cancer

description: Additional file 1:Figure S1. Detailed views of selected pathways identified by WebGestalt analyses. Related to Fig. 3. All panels are adapted from File S6. Heat map colors follow Fig. 3. (A) Principal component (PC) analysis of SWATH-MS proteomics results showing distribution of PC1 and PC2. PC1 corresponded to pathways associated with ribosomes and translation, while PC2 corresponded to pathways associated with mRNA splicing processing (see File S3). (B) Proteins associated with PI3K/AKT activity (WebGestalt Database: PC, DB_ID:1648, “Class I PI3K signaling events mediated by AKT”) are less abundant in TM cells. (C) F1/F0 ATPase subunits elevated in WT and TM cells. (D) Abundances of proteins associated with protein folding and microtubule function are altered by PGRMC1 phosphorylation status. Proteins detected in any of the following WebGestalt pathways or functions (1–4) or a manual search (5) are mapped against their expression profiles. 1) cellular component chaperonin-containing T-complex GO:0005832. 2) PC pathway Chaperonin-mediated protein folding DB_ID:710. 3) cellular component microtubule GO:0005874. 4) PC pathway Protein folding DB_ID:712. 5) Description from the list of 243 proteins (File S6) contains keywords “tubulin” or “microtubule” (manual search) (Adapted from File S6). 2-tailed t-test p-values for all sample comparisons are available in File S4. (E) Proteins associated with nuclear import/export that are elevated in DM cells. (F) Antigen processing and presentation enzymes are affected by PGRMC1 phosphorylation status. Manual additions to KEGG pathway ID:04612 “Antigen processing and presentation" (no yellow shading: from File S6 and File S5) are indicated with yellow highlighting. Figure S2. Highest and lowest differentially abundant proteins. Panels show the six most (+) and least (−) abundant proteins for each cell type that were significantly differentially abundant between cell types. Related to Fig. 3. Identical colored symbols depict the same protein in different cell types, where circles represent high abundance and triangles represent low abundance. E.g. Mitochondrial import receptor subunit TOM40 (O96008), CDP-diacylglycerol-inositol 3-phosphatidyltransferase (CDIPT, O14735) (phosphatidylinositol synthesis) and transcriptional coactivator PSIP1 (O75475) are more abundant in WT and TM. Aldehyde dehydrogenase 1A3 (P47895), APOC3 (P02656) and APOA1 (P02647) are less abundant in DM and TM, whereas the receptor tyrosine kinase ephrin type-A receptor 2 (EPHA2, P29317) is among the lowest abundance differential proteins in WT and TM. Protein abundances (measured ion intensities) for all proteins are available in File S2. Figure S3. Proteins associated with predicted Estrogen Receptor Related 1 (ERR1) transcription factor activity are more abundant in WT and TM cells. Related to Fig. 3 and File S6. Data are available via ProteomeXchange with identifier PXD014789. (A) Proteins quantified by SWATH-MS and predicted by WebGestalt Transcription Factor target enrichment analysis (WebGestalt Database: Transcription Target, Name: hsa_TGACCTY_V$ERR1_Q2, ID:DB_ID:2414) to be dependent upon ERR1 transcription (Adapted from File S6). Heat map colors follow Fig. 3. (B) Western blot of shRNA attenuation of ERR1 (top panel) in WT cells, compared to scramble shRNA control. Lower panel: beta actin loading control. (C) ERR1 attenuation by shRNA induces morphological changes to WT cells. (D) Ratio of rounded to other cells as scored for replicate images from each of the cell lines from B. p 10 fluorescent intensity units to the right of the reference line in (A). n = 6 for each cell type, being 6 replicates of MP cells, or duplicate measurements of each of 3 independent lines 1–3 (n = 3 × 2 = 6) for WT, DM and TM cells. White arrows indicate the same differences as in (A). There was significant difference between the means (Kruskal-Wallis Test p