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. 2021 Aug 5;10(8):755.
doi: 10.3390/biology10080755.

Transcriptome and Literature Mining Highlight the Differential Expression of ERLIN1 in Immune Cells during Sepsis

Susie S Y Huang  1 , Mohammed Toufiq  1 , Luis R Saraiva  1   2 , Nicholas Van Panhuys  1 , Damien Chaussabel  1 , Mathieu Garand  1
Affiliations

Transcriptome and Literature Mining Highlight the Differential Expression of ERLIN1 in Immune Cells during Sepsis

Susie S Y Huang et al. Biology (Basel). .

Abstract

Sepsis results from the dysregulation of the host immune system. This highly variable disease affects 19 million people globally, and accounts for 5 million deaths annually. In transcriptomic datasets curated from public repositories, we observed a consistent upregulation (3.26-5.29 fold) of ERLIN1-a gene coding for an ER membrane prohibitin and a regulator of inositol 1, 4, 5-trisphosphate receptors and sterol regulatory element-binding proteins-under septic conditions in healthy neutrophils, monocytes, and whole blood. In vitro expression of the ERLIN1 gene and proteins was measured by stimulating the whole blood of healthy volunteers to a combination of lipopolysaccharide and peptidoglycan. Septic stimulation induced a significant increase in ERLIN1 expression; however, ERLIN1 was differentially expressed among the immune blood cell subsets. ERLIN1 was uniquely increased in whole blood neutrophils, and confirmed in the differentiated HL60 cell line. The scarcity of ERLIN1 in sepsis literature indicates a knowledge gap between the functions of ERLIN1, calcium homeostasis, and cholesterol and fatty acid biosynthesis, and sepsis. In combination with experimental data, we bring forth the hypothesis that ERLIN1 is variably modulated among immune cells in response to cellular perturbations, and has implications for ER functions and/or ER membrane protein components during sepsis.

Keywords: bacteremia; calcium channel; cholesterol biosynthesis; immunometabolism; innate immunity; leukocytes; myeloid cells; neutrophil; sepsis.

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Conflict of interest statement

The authors declare that they have no conflict of interest.

Figures

Figure 1
Figure 1
Abundance of ERLIN1 transcript and proteins in cultured whole blood and HL60 cells in response to LPS/PGN exposure. (A) Experimental designs for in vitro stimulation of cultured whole blood and HL60 cells. Whole blood was incubated with or without (UN) a combination of lipopolysaccharide (LPS, 100 ng/mL) and peptidoglycan (PGN, 5 ug/mL) for 6 h with stimulation prior to total RNA extraction and RT-qPCR quantification. Determination of protein levels by flow cytometry and cholesterol assays was performed after 24-h stimulation. For HL60, cells seeded at 3E06 cells/mL were differentiated for six days prior to stimulation for 6, 12, and 24 h with LPS/PGN, followed by processing for flow cytometry and cholesterol assays as described in the Materials and Methods section. (B) Whole blood from healthy donors (n = 8) after 6 h with LPS/PGN or culture medium (control). ERLIN1 expression was assessed by RT-qPCR and normalized to GAPDH transcript expression. (C) Representative dot plots of ERLIN1 abundance in cultured whole blood immune cells before and after stimulation for 24 h (expressed as % of parent cell population). After culture, cells were harvested in BD FACS Lyse and stored at −80 °C until staining. Immunostaining and flow cytometry were performed as described in the Materials and Methods section. Gates were calibrated using all-minus-one staining mix (Figure S4). The summary of the main findings shows the relative fold change (STIM/UN) of the percentage frequency of the parent subset. (D) Relative abundance of ERLIN1 in HL60 cells at 6, 12, and 24 h, expressed as the fold change in median fluorescence intensity (MFI) for each stimulation time (stimulated/non-stimulated); blue and red histograms represent stimulated and non-stimulated, respectively. * p-value ≤ 0.05. **** p-value < 0.0001.
Figure 2
Figure 2
Knowledge gaps concerning ERLIN1, key biological concepts, and PubMed literature on sepsis, inflammation, and neutrophils demonstrate potential novel roles of ERLIN1 in the pathogenesis of sepsis. (A–C) The available literature used to infer potential roles for ERLIN1 expression in immune cells during sepsis through ER-mediated protein degradation, cholesterol homeostasis, and signaling events via lipid structures and inositol pathways. Results of literature search for ERLIN1 (and aliases) and the terms sepsis, inflammation, and neutrophils in humans are enclosed in black circles; the numbers represent the article count retrieved for each association. The PubMed search results with the key biological concepts (colored circles) and literature on sepsis, inflammation, and neutrophils are depicted with dotted lines of corresponding colors. (D) Functional grouping of the title terms from the PubMed query on ERLIN1. The extracted terms were manually assembled into 20 representative functional groups (main graph), and then further condensed to a final list of five key biological concepts for subsequent search strategies.
Figure 3
Figure 3
Hypothetical model of the roles of ERLIN1 in cellular metabolic regulation during sepsis. Based on the compendium of evidence from public transcriptomic data, scientific literature reports, and our preliminary results, we assembled relevant biological knowledge about the roles of ERLIN1 in calcium and cholesterol regulation. (Panel A) ERLIN1-mediated regulation of intracellular Ca2+ during sepsis: (1) Toll-like receptor 4 (TLR4) is activated by its ligand l LPS; (2) Second messengers, inositol triphosphate (IP3), and diacylglycerol (DAG) are produced via the enzymatic activity of phospholipase-c (PLCγ); (3) increase in intracellular Ca2+ due to activation of the transient receptor potential ion channel TRPC6 by DAG, and release of ER pool via the binding of IP3 to the inositol triphosphate receptor (IP3R); (4) immediately upon IP3R activation, ERLIN dimer recruits E3 ligase RNF170 [9], and (5) tags the receptor for degradation via ERAD. (Panel B) Putative alteration of cholesterol homeostasis during sepsis: (1) ERLIN1 retains cholesterol (CHOL) and the SREBP–SCAP–Insig complex in the ER membrane [7]; (2) SCAP-induced conformation changes during CHOL insufficiency and leads to dissociation of SREBP from ERLIN1. Independently, CHOL is transported to the plasma membrane via the sterol homeostasis protein (ARV1); (3) ERLIN1 tags Insig-1 for degradation, and the SREBP–SCAP complex is released to the Golgi to activate CHOL synthesis. Septic stress could induce ERLIN1 expression to increase ERAD efficiency. Collaterally, this could promote the retention of intracellular cholesterol. Low levels (below 25.1 mg/dL) of high-density lipoprotein cholesterol (HDL-C) have been strongly associated with the risk of adverse outcomes in sepsis [8].
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