Phospholipid Scramblase 1 Binds to the Promoter Region of the Inositol 1,4,5-Triphosphate Receptor Type 1 Gene to Enhance Its Expression

Abstract

Phospholipid scramblase 1 (PLSCR1) is a multiply palmitoylated, endofacial membrane protein originally identified based on its capacity to promote accelerated transbilayer phospholipid movement in response to Ca 2؉ . Recent evidence suggests that this protein also participates in cell response to various growth factors and cytokines, influencing myeloid differentiation, tumor growth, and the antiviral activity of interferon. Whereas plasma membrane PLSCR1 was shown to be required for normal recruitment and activation of Src kinase by stimulated cell surface growth factor receptors, PLSCR1 was also found to traffic into the nucleus and to tightly bind to genomic DNA, suggesting a possible additional nuclear function. We now report evidence that PLSCR1 directly binds to the 5-promoter region of the inositol 1,4,5-triphosphate receptor type 1 gene (IP3R1) to enhance expression of the receptor. Probing a CpG island genomic library with PLSCR1 as bait identified four clones with avidity for PLSCR1, including a 191-bp fragment of the IP3R1 promoter. Using electrophoretic mobility shift and transcription reporter assays, the PLSCR1-binding site in IP3R1 was mapped to residues ؊101 GTAACCATGTGGA ؊89 , and the segment spanning Met 86 -Glu 118 in PLSCR1 was identified to mediate its transcriptional activity. The significance of this interaction between PLSCR1 and IP3R1 in situ was confirmed by comparing levels of IP3R1 mRNA and protein in matched cells that either expressed or were deficient in PLSCR1. These data suggest that in addition to its role at the plasma membrane, effects of PLSCR1 on cell proliferative and maturational responses may also relate to alterations in expression of cellular IP3 receptors. The phospholipid scramblase (PLSCR)2 gene family consists of an apparent tetrad of genes with identifiable orthologues conserved from Caenorhabditis elegans to man (1). The first identified member of this family (PLSCR1) was isolated based upon its capacity to promote Ca 2ϩ -dependent accelerated transbilayer membrane phospholipid (PL) movement, mimicking the remodeling of plasma membrane PL that is observed under conditions of injury and apoptosis (2, 3). PLSCR1 is a multiply palmitoylated, Ca 2ϩ -binding, Pro-and Cys-rich, endofacial plasma membrane protein that was shown to distribute into lipid raft domains and to be a substrate of the Abl and Src tyrosine kinases (4 -6). The exact biologic function of this protein remains controversial; although PLSCR1 mediates Ca 2ϩ -dependent transbilayer movement of PL in proteoliposomes (2, 3) and was reported to increase cell surface expression of phosphatidylserine through remodeling of plasma membrane PL in mammalian cells exposed to Ca 2ϩ ionophore and other inducers of injury or apoptosis (7-9), it has also been observed that induced elevation of PLSCR1 expression can occur without a detectable increase in PL movement between membrane leaflets, and gene deletion of PLSCR1 did not impair cell capacity to undergo this remodeling of cell surface PL (10 -12). Whereas its ro...

Wogonoside induces cell cycle arrest and differentiation by affecting expression and subcellular localization of PLSCR1 in AML cells

“…34 IP3R1 is known to play a key role in IP3-mediated mobilization of intracellular calcium ion Ca21 stores from the endoplasmic reticulum in a variety of cells and tissues, and it is also essential for cell growth and differentiation. 44,45 IP3R1 expression in many cells is upregulated in response to ATRA, DMSO, and interleukin-1b.…”

“…PLSCR1 was previously reported to traffic into the nucleus and bind to a specific segment of the IP3R1 59-promoter and enhance transcription of this gene. 34 We therefore investigated the promotion of PLSCR1 binding to the IP3R1 gene promoter by wogonoside using EMSA. PLSCR1 binding to double-stranded oligonucleotides containing the IP3R1 promoter was upregulated after treatment with 150 mM wogonoside for 48 hours, while untreated cells showed little or no DNA binding in the promoter ( Figure 5C).…”

Phospholipid Scramblase 1 Is Secreted by a Lipid Raft-dependent Pathway and Interacts with the Extracellular Matrix Protein 1 in the Dermal Epidermal Junction Zone of Human Skin

“…Involvement of PLSCR1 in protein-protein interaction is becoming more evident. It has been proposed that PLSCR1 interacts with several cellular proteins involved in signaling pathways and acts as a transcription factor for the inositol 1,4,5-triphosphate receptor type 1 gene (23,53). In this study, we identified ECM1a as a novel protein interaction partner of PLSCR1 in human skin.…”

“…In summary, although PLSCR1 is a member of the PLSCR gene family of type II transmembrane endofacial plasma membrane proteins and is thought to contribute to the translayer movement of phosphatidylserine ("scrambling") and other membrane phospholipids, the function of PLSCR1 as a scramblase has been challenged (23,41,46,(51)(52)(53). From recent studies, it is becoming clear that the proteins labeled as "phospholipid scramblases" are not required for scramblase activation or activity (for an update on PLSCR, consult Ref.…”

Natural products against hematological malignancies and identification of their targets

“…101 GTAACCATG-TGGA 89 ), enhancing the transcription of this gene [104]. IP3R1 is known to play a key role in IP3-mediated mobilization of intracellular Ca 2+ stores from the ER in a variety of cells and tissues, and it is also essential for cell growth and differentiation [105,106].…”