International Mammalian Genome Society


The 13th International Mouse Genome Conference
October 31-November 3, 1999

Table of Contents * Structure * Bioinformatics * Sequence * Mapping * New Tools * Gene Discovery * Developmental * Mutagenesis * Functional Genomics

F26 Altered Expression of Pten is an Essential Event in Mouse Thymic Lymphomagenesis

Michel Herranz, Javier Santos, Mónica Fernández, Janet Reyes, Concepción Vaquero, and José Fernández-Piqueras. Dpto. Biología. Fac. Ciencias. Universidad Autónoma de Madrid. 28049-Madrid. Spain

In the present study, we have carried out an extension of the genome LOH analysis in a total of 77 g-radiation-induced thymic lymphomas (stage III of the disease) of (C57BL/6J x BALB/cJ) F1 mice (52 tumors) and the reciprocal strain (BALB/cJ x C57BL/6J) F1 (25 tumors) with four polymorphic microsatellites located on mouse chromosome 19 (D19Mit19, D19Mit40, D19Mit46, and D4Mit100). These markers mapped in a region syntenic with the human chromosome band 10q23, which is frequently deleted in wide variety of human tumors and where the tumor suppressor PTEN gene was recently located. LOH was found in 27 of 77 (35.1%) tumors in at least one of the analyzed markers. A minimum region of allelic loss of about 1cM defined by the markers D19Mit46 and D19mit40 as the likely domain of a new tumor suppressor gene, and we named as TLSR6. Interestingly, the map position of mouse Pten lies within the interval defined by the D19Mit46 and D19Mit40 markers. Thus, Pten could be the candidate tumor suppressor gene for TLSR6.

To evaluate its possible involvement in the development of mouse primary thymic lymphomas induced by g-rays we examined the Pten gene for altered expression and for mutation. Previously, we demonstrated that no processed Pten pseudogene exists in the mouse genome. The functional status of Pten was studied, initially, by reverse transcription followed by PCR amplification (RT-PCR) in the same 77 g-radiation-induced primary thymic lymphomas subjected to LOH analysis. We found that while a 35.1% (27/77) of the tumoral samples showed normal expression of Pten by RT-PCR, altered mRNA levels were present in 64.9% (50/77) of the tumors tested. From these, no transcription of Pten were observed in 36 tumors (72%) while the remaining 14 cases (28%) exhibited low transcript levels of Pten. We next studied the expression level of Pten in the same panel of g-radiation-induced primary thymic lymphomas by Western blot, and found that 65 of 77 (84.4%) of tumors showed variation in the Pten protein levels. Mutation screening by SSCP analysis of the entire Pten coding region showed no mobility shifts in tumor samples as compared with normal thymus. These suggest that altered expression rather than point mutations of the Pten gene seems to play a role in the development of mouse primary thymic lymphomas induced by g-rays. In addition, inactivation of Pten is correlated in a significant fraction of the tumors analyzed with detectable levels of phosphorylated Akt/PKB. Thus, Pten inactivation might facilitate a cell survival pathway via Bcl2.

Mouse thymic lymphomas induced by g-rays are an excellent model for studying tumor progression, since the development of this type of cancer is defined by two clearly distinguishable steps: the early disease stages (I and II) and the frank lymphomas (stage III). To investigate the moment in tumor development in which Pten inactivation occur, we have examined 30 samples in early disease stages, taken within 12 weeks (15 samples) and 27 weeks (15 samples) following the g-irradiation treatment, for altered expression of Pten. While abnormalities in Pten expression were not detected in any of samples taken within 27 weeks, 5 of 15 (33.3%) samples in an early disease stage within 12 weeks showed altered RNA or protein levels of Pten. These findings suggest that Pten inactivation appears to play an important role in tumor initiation rather than in progression.

 


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