Leukemia is one of the most deadly diseases, responsible for the highest number of childhood cancer cases. Immunoglobulin-containing and proline-rich receptor-1 (IGPR-1) is a newly identified protein found to play an important role in human colon cancer and angiogenesis. The overall goal of this project was to assess IGPR-1 expression in leukemia cell lines and investigate its possible function in the NF-κB pathway, specifically the role of inhibitor of nuclear factor kappa-B kinase subunit beta (IKKβ) in the phosphorylation of IGPR-1. The NF-κB pathway, among others, plays a critical role in human-T-cell leukemia virus type 1 (HTLV-1) infected T-cells. Our preliminary results indicated that IGPR-1 is expressed in leukemia cell lines at variable levels. Further experiments demonstrated that IKKβ is involved in the phosphorylation of IGPR-1 as treatment of HEK-293 cells ectopically expressing IGPR-1 with an IKKβ inhibitor decreased IGPR-1 phosphorylation at Ser220. Likewise, cells treated with lipopolysaccharide (LPS), which is known to activate IKKβ, also stimulated the phosphorylation of IGPR-1 at Ser220. However, transfection of IGPR-1/HEK293 cells with Tax, an oncogene encoded by HTLV-1, decreased phosphorylation of IGPR-1 at Ser220. Taken together, our data indicates that activation of IKKβ in the NF-κB pathway stimulates phosphorylation of IGPR-1. / 2020-06-17T00:00:00Z

Immunoglobulin-containing and proline-rich receptor-1 (IGPR-1), is a newly identified cell adhesion molecule and is expressed in various cell types including, epithelial and endothelial cell origin. IGPR-1 regulates cell-cell adhesion and promotes angiogenesis, activated by shear stress and mediates endothelial cells response to shear stress. Moreover, IGPR-1 expression is upregulated in colon cancer and supports colon tumor growth. IGPR-1 contains a single extracellular immunoglobulin domain, a transmembrane domain and followed by a cytoplasmic proline-rich C-terminus. We demonstrate that ubiquitin E3 protein ligase neural precursor cell expressed developmentally down regulated 4 (NEDD4) binds to and ubiquitinates IGPR-1. Furthermore, among the four WW domains, the C-terminus WW domain#4 selectively mediates the binding of NEDD4 with IGPR-1. We used in vitro ubiquitination assay and identified UbcH6, as an E2 conjugating enzyme required for NEDD4-mediated ubiquitination of IGPR-1. Taken together, our data identifies NEDD4/Ubc6H ubiquitination system as a major pathway involved in the ubiquitination of IGPR-1. / 2021-06-08T00:00:00Z

Disruption of the integrity of vascular endothelium plays an essential role in the development and the progression of numerous human diseases, including sepsis, atherosclerosis and others. A complex array of transmembrane adhesive proteins located in junctional structures, support endothelial integrity and control vascular permeability. Furthermore, they are able to transmit intracellular signals to coordinate various endothelial biological responses to insure normal vascular function. Immunoglobulin-containing and proline rich receptor-1 (IGPR-1) is a novel cell adhesion molecule that is involved in angiogenesis and in the regulation of endothelial permeability. IGPR-1 is phosphorylated at Ser220, which is required for its ability to mediate actin fibril reorganization. In this study, we demonstrate that the phosphorylation of IGPR-1 at Ser220 is stimulated by cell spreading and cell adhesion in porcine aortic endothelial (PAE) cells. Blocking homophilic trans-dimerization of IGPR-1 by a blocking antibody inhibited cell-density phosphorylation of IGPR-1. More importantly, phosphorylation of IGPR-1 at Ser220 is increased in PAE cells under shear stress, which was essential for IGPR-1-mediated endothelial cell alignment in response to shear stress. Taken together, this study demonstrate that IGPR-1 activity is regulated be endothelial cell spreading and density. And its activity plays an important role in endothelial cell alignment in response to shear stress. / 2020-07-03T00:00:00Z

Colorectal cancer (CRC) is among the most prevalent and lethal cancers in the United States. The mechanisms by which tumor cells sense their microenvironment have profound importance in driving the progression of malignancy and evasion from treatment. Specialized microenvironment-sensing cell surface receptors such as cell adhesion molecules allow tumor cells to survey and respond to their microenvironment. We have recently identified a novel cell adhesion molecule named immunoglobulin-containing and proline-rich receptor 1 (IGPR-1) that is normally expressed in both endothelial and epithelial human cell types; however, its potential role in human malignancy remains unknown. To investigate the role IGPR-1 plays in CRC tumor growth, we overexpressed IGPR-1 in human HT29 and HCT116 colon adenocarcinoma cells and examined the effect of IGPR-1 on tumor growth and the mechanisms involved in a cell culture system. The data demonstrate that overexpression of IGPR-1 enhances CRC cell proliferation and survival in vitro. Furthermore, we demonstrate that the extracellular domain of IGPR-1 is required for its ability to support tumor growth. While deletion of the extracellular domain of IGPR-1 impaired its ability to promote tumor cell survival, stimulation of the chimeric IGPR-1 consisting of the extracellular domain of human colony stimulating factor-1 receptor (CSF-1R) fused to the transmembrane and cytoplasmic domains of IGPR-1 promoted tumor cell survival. Additionally, the presence of serine 186 and 220 in the cytoplasmic domain is important for IGPR-1 activity in tumor cells. This work identifies IGPR-1 as an important protein in the regulation of CRC cell growth and survival, and this makes it a possible therapeutic target in the clinical management of CRC.

Angiogenesis is a physiological process by which new blood vessels develop from preexisting vasculature. The process of converting endothelial cells into fully developed blood vessels involves multiple coordinated cellular events that occur through the collaboration that exists between a variety of growth factors, receptors and adhesion molecules. The immunoglobulin-containing and proline rich receptor-1 (IGPR-1) is an IgSF containing adhesion molecule that has been recently identified as a novel regulator of angiogenesis in vitro. In this study, we provide evidence that IGPR-1 promotes cell survival in porcine aortic endothelial cells (PAE) and plays a role in the inhibition of p38 MAPK in vitro. Deletion of the extracellular domain of IGPR-1 abolished IGPR-1’s ability to inhibit phosphorylation of p38 MAPK and promote the survival of endothelial cells. Likewise, mutation of serines 186 (A186-IGPR-1) and 220 (A220-IGPR-1) on the cytoplasmic domain of IGPR-1 was also found to reduce both the promotion of cell survival and inhibition of p38 MAPK. These findings suggest that both domains of IGPR-1 are important for endothelial cell survival and the activation p38 MAPK.

Colorectal cancer (CRC) is the third leading cause of cancer-related death in women and fourth in men globally. While expansions in preventative measures have increased the detection of CRC at the early stages of disease, only 40% of CRC patients are diagnosed when the disease is at a local stage. Moreover, many anti-cancer drugs fail to significantly improve the life expectancy of patients due to innate and acquired resistance, underscoring a need for better diagnostic and therapeutic strategies for CRC. Immunoglobulin-containing and proline-rich receptor-1 (IGPR-1) is a novel cell adhesion molecule (CAM) that was recently identified in our laboratory. IGPR-1 is expressed in epithelial and endothelial cells and promotes cell-cell adhesion. Expression of IGPR-1 in endothelial cells regulates angiogenesis; however, its role in epithelial cells, particularly cancer cells with an epithelial origin, remains unknown. The overall goal of this study was to investigate the possible function of IGPR-1 in CRC tumor cell growth and response to chemotherapeutic agents. Specifically, we aimed to test the hypothesis that increased expression of IGPR-1 in CRC tumor cells promotes cell survival and contributes to the resistance of tumor cells to doxorubicin. Human CRC tumor cell lines, HCT116 and HT29, were transduced via a retroviral system to express IGPR-1 or empty retroviral vector pQCXIP. The effect of overexpression of IGPR-1 in HCT116 and HT29 cells was measured by MTT assay in non-adherent 24-well plates. In addition, cells were viewed under a light microscope, and images were taken to assess multicellular aggregation. Results demonstrated that expression of IGPR-1 in HCT116 and HT29 tumor cells promoted CRC tumor cell growth, increased multicellular aggregation, and stimulated resistance to the conventional chemotherapeutic agent doxorubicin in non-adherent cell culture conditions in vitro. Intriguingly, treatment of cells with doxorubicin promoted phosphorylation of IGPR-1 at serine 220 (Ser220), suggesting a critical role for phosphorylation of IGPR-1 in the development of resistance to chemotherapeutics. In addition, non-adherent cell culture conditions promoted activation of the key pro-apoptotic kinase, p38 MAPK in CRC tumor cells. Ectopic expression of IGPR-1 reversed this activation. This data suggests that IGPR-1, by suppressing p38 activity, in part, promotes tumor cell survival and increases the resistance of tumor cells to the killing effects of doxorubicin. Our findings are the first to demonstrate that IGPR-1 promotes CRC tumor cell growth and increases the resistance of CRC tumor cells to the cytotoxic effects of chemotherapeutic agents. The data suggests that IGPR-1 plays an important role in CRC by inhibiting the cellular apoptotic response and promoting chemotherapeutic resistance. Finally, IGPR-1 phosphorylation at Ser220 in response to doxorubicin may account for the IGPR-1-mediated development of resistance to doxorubicin in CRC.

Dysregulation of endothelial cell barrier function is associated with a wide variety of human diseases ranging from tumor metastasis to inflammation. The barrier function of endothelial cells is maintained by cell adhesion molecules (CAMs). Immunoglobulin containing and proline-rich receptor-1(IGPR-1) was recently identified as a novel CAM involved in angiogenesis. However, the molecular mechanism of IGPR-1 function in endothelial cells remains largely unknown. The overarching goals of this study were: (A) to determine molecular mechanism by which IGPR-1 stimulates biological responses in cells and (B) to investigate regulation of phosphorylation of IGPR-1 at serine 220 (Ser220), and its role in IGPR-1 function. Our data demonstrate that IGPR-1 undergoes cis-dimerization, which leads to homophilic trans-dimerization of IGPR-1, which is required for its adhesive function. Moreover, we demonstrate that phosphorylation of Ser220 is regulated by trans-dimerization of IGPR-1 and that Glycogen Synthase Kinase 3 (GSK-3) is responsible for its phosphorylation as over-expression of kinase active increased and kinase inactive inhibited phosphorylation of Ser220, respectively. Taken together, the results demonstrate that the coordinated dimerization of IGPR-1 and its homophilic interaction regulates its adhesive function and serine phosphorylation. The adhesive function of IGPR-1 contributes to the barrier function of endothelial cells.