Abnormal splicing of SHP-1 protein tyrosine phosphatase in human T cells. Implications for lymphomagenesis.
OBJECTIVE
SHP-1 protein tyrosine phosphatase has been implicated in suppressing B-lymphocyte and myeloid cell malignancies; however, there are little data on this role of SHP-1 in T-lymphocyte malignancies. We examined malignant human T cells to identify any abnormalities of SHP-1 that would support a role for this molecule in suppressing T lymphomagenesis.


METHODS
Human T-lymphocyte cell lines and primary blood cells were used to examine the expression of SHP-1 mRNA and protein. Reverse transcriptase polymerase chain reaction was used to amplify particular portions of the SHP-1 mRNA for cloning and sequencing. Gene transfer was used to examine the effects of SHP-1 on cell growth and morphology. Glutathione S-transferase (GST) fusion proteins were generated and used to determine SHP-1-associated proteins.


RESULTS
Leukemia- and lymphoma-derived cell lines were identified that did not express SHP-1 protein. Examination of the mRNA from these and other T-cell lines, and from normal peripheral blood mononuclear cells (PBMCs), revealed three distinct transcripts by restriction enzymes, reverse transcriptase polymerase chain reaction, and Southern blot analysis. In addition to the expected wild-type transcript, two novel transcripts were identified. One was a deletion transcript found only in Jurkat leukemia-derived cells, predicted to encode for a 7-kDa protein containing most of the amino-terminal SH2 domain. The second contained an 88-nucleotide insert that is the unspliced second intron resulting in a frame shift and the formation of a noncoding transcript. This mRNA was found in all cells examined but was the only transcript detected in the cell lines lacking SHP-1 protein. Expressing wild-type SHP-1 in these cell lines resulted in a change in the morphology of the cells with a concomitant decrease in their growth. GST fusion constructs showed the 7-kDa variant able to associate with an identical array of proteins as wild-type SHP-1, suggesting that it could compete with the wild-type SHP-1 for substrates. This protein was detectable in the cell line expressing its corresponding mRNA and was able to induce significant changes in cell morphology when transfected into a cell line expressing wild-type SHP-1; however, it did not induce any changes in cell growth.


CONCLUSIONS
These data are the first to show the existence of multiple transcripts of SHP-1 in human transformed T lymphocytes and normal PBMCs and supports previous work showing that alternate forms of SHP-1 mRNA are a common finding in other cells. We also show the lack of splicing out of an intron as a novel mechanism of regulation of SHP-1 protein expression in both normal and transformed T cells. Moreover, we provide the first evidence showing a protein product detectable in cells that is translated from an alternatively spliced form of SHP-1 mRNA, a variant truncated SHP-1 protein having potential biologic relevance. This report provides evidence supporting the concept that SHP-1 can negatively regulate growth of malignant human T cells and that lack of SHP-1 protein or function may be associated with lymphomagenesis.
Ma
Xue-Zhong
XZ
Department of Medicine and Institute of Medical Science, University of Toronto, 67 College Street, Toronto M5G 2M1, Ontario, Canada
Jin
Tianru
T
Sakac
Darinka
D
Fahim
Soad
S
Zhang
Xin
X
Katsman
Yulia
Y
Bali
Meenakshi
M
Branch
Donald R
DR
D017398:Alternative Splicing / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D002455:Cell Division / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007938:Leukemia / Q000201:enzymology / Q000209:etiology; D008213:Lymphocyte Activation / Q000187:drug effects; D008223:Lymphoma / Q000201:enzymology* / Q000209:etiology; D008969:Molecular Sequence Data; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000235:genetics* / Q000494:pharmacology / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics; D017423:Sequence Analysis, RNA; D013601:T-Lymphocytes / Q000201:enzymology* / Q000473:pathology; D014162:Transfection; D014407:Tumor Cells, Cultured
D047908:Intracellular Signaling Peptides and Proteins; D012333:RNA, Messenger; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases