Detection and analysis of an alternatively spliced isoform of interleukin- 6 mRNA in peripheral blood mononuclear cells

Daniel Kestler, S. Agarwal, J. Cobb, K. M. Goldstein, R. E. Hall

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Oligonucleotide primers for human interleukin-6 (IL-6) that bracketed the entire coding region of the gene were used in reverse transcriptase- polymerase chain reaction (RT-PCR) studies to examine IL-6 expression in peripheral blood mononuclear cells (PBMC). In addition to the predicted 0.64- kb RT-PCR product, a second 0.45-kb product was observed. Cloning and dideoxy sequence analysis of this product revealed evidence for an alternatively spliced IL-6 transcript lacking exon II. Further RT-PCR analysis using forward primers ending at or one base before the exon I donor splice site again yielded both products. Additional primers were designed and successfully used to selectively distinguish the two forms of IL-6 transcript. Both transcripts were prominent in peripheral blood monocytes and lymphocytes, whereas only the 0.64-kb, full-length transcript was prominent in the IL-6-producing 5637 (human bladder carcinoma) cell line. Northern analysis revealed, in addition to the predominant 1.3-kb transcript, several minor transcripts at 1.9 to 4.8 kb that hybridized with the alternatively spliced cDNA probe but not with an exon II probe. Western analysis revealed IL-6 polypeptides of predicted size (26 to 29 kD) in culture medium from PBMC, while showing an immunoreactive band at 17 kD in cell lysates. These findings suggest the existence of an alternatively spliced form of IL-6 mRNA, which would encode for a polypeptide missing the gp130 interactive (signal- transducing) domain contained in exon II while retaining the IL-6 receptor (p80) domain. Such a molecule could in theory function as a natural antagonist of IL-6, as it would be expected to bind to the IL-6 receptor but not lead to signal transduction.

Original languageEnglish (US)
Pages (from-to)4559-4567
Number of pages9
JournalBlood
Volume86
Issue number12
StatePublished - Jan 1 1995
Externally publishedYes

Fingerprint

Interleukin-6
Blood Cells
Protein Isoforms
Blood
Messenger RNA
Exons
Polymerase chain reaction
RNA-Directed DNA Polymerase
Reverse Transcriptase Polymerase Chain Reaction
Interleukin-6 Receptors
Signal transduction
Peptides
RNA Splice Sites
DNA Primers
Lymphocytes
Cloning
Reaction products
Sequence Analysis
Culture Media
Organism Cloning

All Science Journal Classification (ASJC) codes

  • Hematology

Cite this

Detection and analysis of an alternatively spliced isoform of interleukin- 6 mRNA in peripheral blood mononuclear cells. / Kestler, Daniel; Agarwal, S.; Cobb, J.; Goldstein, K. M.; Hall, R. E.

In: Blood, Vol. 86, No. 12, 01.01.1995, p. 4559-4567.

Research output: Contribution to journalArticle

Kestler, D, Agarwal, S, Cobb, J, Goldstein, KM & Hall, RE 1995, 'Detection and analysis of an alternatively spliced isoform of interleukin- 6 mRNA in peripheral blood mononuclear cells', Blood, vol. 86, no. 12, pp. 4559-4567.
Kestler, Daniel ; Agarwal, S. ; Cobb, J. ; Goldstein, K. M. ; Hall, R. E. / Detection and analysis of an alternatively spliced isoform of interleukin- 6 mRNA in peripheral blood mononuclear cells. In: Blood. 1995 ; Vol. 86, No. 12. pp. 4559-4567.
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