The role of CD150-SH2D1A association in CD150 signaling in Hodgkin's lymphoma cell lines.
OBJECTIVE
To find out what signal transduction pathways are linked to CD150 in Hodgkin's lymphoma (HL) cell lines, how are they regulated, and to examine the expression of different CD150 splice forms and SH2 domain containing protein D1A (SH2D1A) adaptor protein on mRNA level in HD primary tumors and cell lines.


METHODS
The expression of CD150 splicing forms and SH2D1A adaptor protein in HD primary lymphoma tissue and cell lines were analyzed by RT-PCR method. To examine CD150-SH2D1A localization in HD cell lines we performed double immunofluorescent staining of these two proteins. Total amount of SH2D1A, Syk, Lyn, SHP-2, SHIP proteins, and activated/phosphorylated ERK1/2 and Akt proteins were detected by Western blot analysis with specific antibodies.


RESULTS
We showed the expression of soluble (sCD150) and full length transmembrane (mCD150) splice forms and SH2D1A adaptor protein on mRNA level in 9 cases of classical HD and three HD lines of B cell origin - L428, KM-H2 and L1236. In spite of CD150 and SH2D1A co-expression in studied HD cell lines, CD150 co-precipitated and co-localized with SH2D1A only in L1236 cells. CD150 ligation induced extracellular signal-regulated kinases (ERK) dephosphorylation in L1236 cell line, but had no effect on ERK pathway in KM-H2 and L428 cells. CD150 crosslinking induced Akt activation also only in L1236 cells.


CONCLUSIONS
HD cells express sCD150 and mCD150 splice forms and SH2D1A. Association of CD150 and SH2D1A depends at least on their localization pattern. CD150 is linked to ERK and Akt pathways in HD cell lines. Our data suggest that CD150-SH2D1A association play decisive role in Akt signaling upon CD150 ligation in HD cell lines. CD150-mediated Akt activation in HD cell lines, similarly to DT40 model system, is SHIP-independent.
Yurchenko
Maria Y
MY
R.E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology NAS Ukraine, Signal Transduction Laboratory, Kiev, Ukraine. myurchenko@hotmail.com
Kashuba
Elena V
EV
Shlapatska
Larysa M
LM
Sivkovich
Svetlana A
SA
Sidorenko
Svetlana P
SP
D015703:Antigens, CD; D001483:Base Sequence; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins / Q000378:metabolism*; D006689:Hodgkin Disease / Q000378:metabolism*; D006801:Humans; D007136:Immunoglobulins / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface; D020133:Reverse Transcriptase Polymerase Chain Reaction; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D015398:Signal Transduction / Q000502:physiology*; D000071179:Signaling Lymphocytic Activation Molecule Associated Protein; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1; D000072377:Syk Kinase; D019061:src-Family Kinases / Q000378:metabolism
D015703:Antigens, CD; D004792:Enzyme Precursors; D006023:Glycoproteins; D007136:Immunoglobulins; D047908:Intracellular Signaling Peptides and Proteins; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C114953:SH2D1A protein, human; C000604059:SLAMF1 protein, human; D000071179:Signaling Lymphocytic Activation Molecule Associated Protein; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D010744:Phosphoric Monoester Hydrolases; C516724:PTPN11 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; C098706:INPPL1 protein, human; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases