Isolation and characterization of novel RECK tumor suppressor gene splice variants
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Marina Trombetta-Lima1,2, Sheila Maria Brochado Winnischofer3, Marcos Angelo Almeida Demasi1,2, Renato Astorino Filho1,2, Ana Claudia Oliveira Carreira1,2, Beiyang Wei5, Thais de Assis Ribas2, Michelle Silberspitz Konig2, Christian Bowman-Colin2,4, Sueli Mieko Oba-Shinjo2,6, Suely Kazue Nagahashi Marie2,6, William Stetler-Stevenson5, Mari Cleide Sogayar1,2
1Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo 05508-000 SP, Brazil
2NUCEL-NETCEM-Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP, 05360-120, Brazil
3Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Paraná, Curitiba, PR, 81531-990, Brazil
4Dana Farber Cancer Institute, Harvard Medical School, Cambridge, MA, 02138, USA
5Radiation Oncology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892-4605, USA
6Departmento de Neurologia, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP, 01246-000, Brazil
Mari Cleide Sogayar, e-mail: [email protected]
Keywords: RECK, GBM, splicing, isoforms, MMP
Received: March 18, 2015 Accepted: September 15, 2015 Published: September 28, 2015
Glioblastoma multiforme is the most common and lethal of the central nervous system glial-derived tumors. RECK suppresses tumor invasion by negatively regulating at least three members of the matrix metalloproteinase family: MMP-9, MMP-2, and MT1-MMP. A positive correlation has been observed between the abundance of RECK expression in tumor samples and a more favorable prognosis for patients with several types of tumors. In the present study, novel alternatively spliced variants of the RECK gene: RECK-B and RECK-I were isolated by RT-PCR and sequenced. The expression levels and profiles of these alternative RECK transcripts, as well as canonical RECK were determined in tissue samples of malignant astrocytomas of different grades and in a normal tissue RNA panel by qRT-PCR. Our results show that higher canonical RECK expression, accompanied by a higher canonical to alternative transcript expression ratio, positively correlates with higher overall survival rate after chemotherapeutic treatment of GBM patients. U87MG and T98G cells over-expressing the RECK-B alternative variant display higher anchorage-independent clonal growth and do not display modulation of, respectively, MMP-2 and MMP-9 expression. Our findings suggest that RECK transcript variants might have opposite roles in GBM biology and the ratio of their expression levels may be informative for the prognostic outcome of GBM patients.
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