Isolation and characterization of NK cell or NK/T cell-specific cDNA clones.
Natural killer (NK) cells are cytotoxic lymphocytes that share numerous cell surface antigens and functional components with T cells. However, in comparison with our knowledge of T cells, little is known about the molecular mechanisms of NK cell activation and function. The following study was initiated as an effort to obtain further information about similarities and differences between NK and T cells at the level of gene expression and also to identify NK-specific cDNA clones for future functional studies of the corresponding gene products. The study used cDNA libraries prepared from an NK clone and from an Epstein-Barr virus transformed B cell lymphoblastoid cell line (LCL). We employed a combination of differential and subtractive hybridization methodologies, which can successfully identify cell-specific cDNA clones representing medium to high abundance transcripts, to identify genes that are expressed in NK cells but not in the LCL. We were particularly interested to ascertain to what extent genes isolated in this manner would be expressed only in NK cells as opposed to being expressed in NK and T cells. Twelve different cross-hybridizing groups were identified that were not expressed in the LCL, and these groups were further characterized: (1) they were used to probe Northern blots prepared from a panel of cells including NK cells, T cells, and B cells: (2) changes in the steady-state level of message following T cell growth factor (TCGF)-induced activation of an NK cell clone were examined for selected isolates; and (3) a partial DNA sequence was determined for each cross-hybridizing group. The DNA sequences of seven groups were identical to previously reported sequences. One group was highly homologous with but not identical to what has been reported as a T cell specific gene, named 519. The DNA sequences of four groups showed no significant homology with the sequences in the GenBank and EMBL databases. The mRNA expression of the newly-identified groups demonstrated several different regulation patterns with respect to cell distribution and level of expression in response to TCGF-activation. Expression of the twelve different genes was examined in three populations of NK cells all of which were CD3- and possessed NK activity. Although these cells differentially expressed the prototype NK markers CD16 and CD56 (the cells were CD16+, CD56-, CD16-, CD56+ and CD16+, CD56+), the expression of all groups of cDNA clones was comparable in the three different types of NK cells despite the phenotypic differences.(ABSTRACT TRUNCATED AT 400 WORDS)
Houchins
J P
JP
Dept. of Laboratory Medicine, University of Minnesota, Minneapolis 55455.
Yabe
T
T
McSherry
C
C
Miyokawa
N
N
Bach
F H
FH
D015152:Blotting, Northern; D002461:Cell Line, Transformed; D002999:Clone Cells; D003001:Cloning, Molecular; D004247:DNA / Q000032:analysis*; D015342:DNA Probes; D015870:Gene Expression; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D007694:Killer Cells, Natural / Q000737:chemistry*; D008213:Lymphocyte Activation / Q000187:drug effects / Q000235:genetics; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000302:isolation & purification; D013601:T-Lymphocytes / Q000737:chemistry*
D015342:DNA Probes; D007376:Interleukin-2; D012333:RNA, Messenger; D004247:DNA