SV40 Tag transformation of the normal invasive trophoblast results in a premalignant phenotype. II. Changes in gap junctional intercellular communication.
Poor gap junctional intercellular communication (GJIC) has been associated with uncontrolled cell growth and neoplasia. We have successfully propagated normal first trimester invasive extravillous trophoblast (EVT) cells, and have produced premalignant EVT lines after SV40 Tag transformation: RSVT-2 is an uncloned line that is long-lived; RSVT2/C is a clonal line that is immortal. Both are hyperproliferative, hyperinvasive and variably refractory to the anti-proliferative and anti-invasive effects of transforming growth factor beta (TGFbeta). Possible changes in gap junctions during the transition of normal invasive EVT cells to the premalignant stage were examined by comparing expression of connexin proteins (by immunolabeling for Cx26, Cx32, Cx40, Cx43), and mRNA (by Northern blot with cDNA probes for Cx26, Cx32, Cx43), and functional GJIC (by dye transfer using the preloading method) in normal parental EVT cells and their SV40 Tag transformants. Results from immunofluorescence and Northern blot analysis revealed that, of the panel of connexins examined, only Cx43 was variably expressed in these cell lines in vitro. Expression of Cx43 protein and mRNA was abundant in normal EVT cell line HTR8, reduced in long-lived RSVT-2 cells and undetectable in immortalized RSVT2/C cells. GJIC, as measured by dye transfer between donor and recipient cells, was also similarly reduced in recipient RSVT-2 cells, and drastically reduced in RSVT2/C cells, irrespective of whether the dye donor was of the same cell type (homocellular coupling) or HTR8 cells (heterocellular coupling). Treatment with TGFbeta reduced Cx43 mRNA expression as well as GJIC in normal EVT cells, but not in the SV40 Tag transformants. Our findings suggest that downregulation of connexins with the resultant impairment in GJIC is an early event in tumor progression, as observed in the premalignant SV40 Tag transformants.
Khoo
N K
NK
Department of Anatomy and Cell Biology, The University of Western Ontario, London, Canada.
Zhang
Y
Y
Bechberger
J F
JF
Bond
S L
SL
Hum
K
K
Lala
P K
PK
D000704:Analysis of Variance; D000957:Antigens, Viral, Tumor / Q000096:biosynthesis / Q000235:genetics; D002450:Cell Communication / Q000502:physiology*; D002471:Cell Transformation, Neoplastic; D000072259:Connexin 26; D018031:Connexin 43 / Q000032:analysis / Q000096:biosynthesis; D017630:Connexins / Q000032:analysis / Q000096:biosynthesis; D005260:Female; D017629:Gap Junctions / Q000473:pathology / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D009361:Neoplasm Invasiveness / Q000473:pathology*; D010641:Phenotype; D011230:Precancerous Conditions / Q000473:pathology*; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D012333:RNA, Messenger / Q000096:biosynthesis; D013539:Simian virus 40 / Q000235:genetics; D014158:Transcription, Genetic; D014327:Trophoblasts / Q000166:cytology / Q000473:pathology*
D000957:Antigens, Viral, Tumor; D018031:Connexin 43; D017630:Connexins; C000607042:GJB2 protein, human; D012333:RNA, Messenger; D000072259:Connexin 26