Cloning and expression of an alternatively spliced mRNA encoding a soluble form of the human interleukin-6 signal transducer gp130.
The membrane-bound gp130 glycoprotein acts as an affinity converting and signal transducing receptor (R) for interleukin-6 and several other cytokines. In this work, we RT-PCR amplified gp130 cDNA using primers flanking the sequence encoding the transmembrane domain of gp130. We observed in blood mononuclear cells, in addition to the expected 333-bp length fragment, a second major band of 418 bp. Sequencing of the 418-bp fragment and its genomic counterpart showed a new 85-bp exon located in the sequence encoding the extracellular region of the gp130 protein. This exon is most likely due to alternative splicing and leads to a frame-shift resulting in a stop-codon 1 bp before the transmembrane coding region. Correspondingly, supernatants from chinese hamster ovary cells transfected with this cDNA contained 4-5 times more soluble (s) gp130 than supernatants from cells transfected with a cDNA encoding the membrane-bound gp130 protein. Both gp130 and alternatively spliced sgp130 were also transcribed by the myeloma cell lines XG-1, XG-2, XG-4, XG-4CNTF XG-6, XG-7, XG-9, XG-10, U266 and RPMI 8226. However, XG-4A cells derived from XG-4 cells, but growing independently of exogenous IL-6, did not transcribe sgp130 mRNA. A possible interference with intracrine stimulatory factors by alternatively spliced sgp130 needs to be further investigated.
Diamant
M
M
Institute for Inflammation Research, Rigshospitalet, Copenhagen N, Denmark. md@rh.dk
Rieneck
K
K
Mechti
N
N
Zhang
X G
XG
Svenson
M
M
Bendtzen
K
K
Klein
B
B
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000235:genetics*; D001483:Base Sequence; D016466:CHO Cells; D002478:Cells, Cultured; D003001:Cloning, Molecular; D006224:Cricetinae; D050822:Cytokine Receptor gp130; D006801:Humans; D015850:Interleukin-6; D008562:Membrane Glycoproteins / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*; D015398:Signal Transduction
D015703:Antigens, CD; C494199:IL6ST protein, human; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D050822:Cytokine Receptor gp130