Expression of transduced tropomyosin 1 cDNA suppresses neoplastic growth of cells transformed by the ras oncogene.

Abstract

Synthesis of certain members of the tropomyosin family of microfilament-associated proteins is suppressed in fibroblasts neoplastically transformed by a number of retroviral oncogenes, by transforming growth factor a, and by chemical mutagens. To test whether tropomyosin suppression is a required event in neoplastic transformation, expression of one of two suppressed tropomyosins in NIH 3T3 mouse cells transformed by the ras oncogene was restored by retrovirally mediated cDNA transfer. Cells expressing the inserted cDNA showed partial restoration of microfflament bundle formation (which is typicafly deranged in transformed cells) together with increased cytoplasmic spreading. More importantly, they lost anchorage-independent growth capability, and the onset of tumor growth in athymic mice was delayed. When tumors arose they no longer expressed the inserted cDNA. These observations support the conclusion that tropomyosin suppression is a necessary event for the expression of components of the transformed phenotype, particularly with respect to anchorage-independent growth and tumorigenesis, which correlate closely with neoplastic potential. This potentially reversible requirement may link different initial events produced by a variety of oncogenic modalities to a common pathway leading to neoplastic growth.Normal cells may be transformed in culture to the neoplastic state by any of a number of retroviral or activated oncogenes alone or in combination (1-4), by transforming growth factors (5, 6), or by chemical mutagens (7,8). Transformation of fibroblasts by many of these modalities is commonly accompanied by reduced synthesis of various actin-binding proteins, of which tropomyosins (TMs) (9-14), vinculin (15), gelsolin (16), and a-actinin (17) are examples. Consistent loss of expression of specific TM isoforms has also been found in cells cultured from human breast cancer (18). TMs and the other proteins named are essential structural and functional components of the actin-microfilament system of the cell (19-24), which has long been known to be severely disrupted in transformed cells (25,26).In the present study, to clarify whether TM suppression is a necessary event in the causal chain leading to neoplastic transformation, we have examined the effect of overcoming the deficiency of TM synthesis in retrovirally transformed cells (DT cells) through expression of a cDNA encoding the TM1 isoform. DT cells are NIH 3T3 mouse fibroblasts transformed by the v-Ki-ras oncogene and contain two integrated copies of the oncogene (27). They are highly tumorigenic in athymic mice, manifest anchorage-independent growth, and rarely produce spontaneous revertants. Of the five major TM isoforms expressed in NIH 3T3 cells, TM1 is suppressed by 50% and TM2 is almost absent in DT cells, whereas expression of the low molecular weight isoforms TM4 and TM5 is unaltered (9, 10). Nontransformed revertants of DT induced by chemical mutagens reexpress TM1The publication costs of this article were defrayed in part by page charge pa...

v-Src suppresses SHPS-1 expression via the Ras-MAP kinase pathway to promote the oncogenic growth of cells

“…Candidate type II tumor suppressor genes such as NO3 (Ozaki and Sakiyama, 1994), maspin (Zou et al, 1994), ela®n (Zhang et al, 1995), a-actinin (Gluck et al, 1993), tropomyosin 1 and 2 (Prasad et al, 1993;Gimona et al, 1996) and SSeCKS (Lin and Gelman, 1997) have been isolated. Although none of their anti-tumor functions has been clari®ed yet, their de®nition as tumor suppressors is strengthened by at least two criteria (Lin and Gelman, 1997).…”

Suppression of src-induced transformed phenotype by expression of tropomyosin-1

“…The src-T1 cells which were The gels were scanned using an AMBIS radioanalytical imaging system and the amount of radioactivity in TM1 was calculated/ 10 6 c.p.m. of total radioactivity present on the gel (Prasad et al, 1993(Prasad et al, , 1994 TM1-mediated tumor suppression GL Prasad et al generated by transduction of 3T3/src cells with TMe1 cDNA, elaborate up to 20-fold higher TM1 protein (Figure 1b and c). This profound enhancement in the TM1 synthesis is due to the expression from the transduced cDNA sequence.…”

“…For example, in ®broblasts, a total of ®ve di erent TMs are expressed: TM1 (M r , 41 kDa), TM2 (M r , 36 kDa) and TM3 (M r , 35 kDa), each with 284 aa, are referred to as high M r isoforms. TM4 and TM5 (each with M r of 32 kDa) contain 247 aa and are described as low M r isoforms (Cooper et al, 1985;Prasad et al, 1993). The function of TMs in muscle cells, in conjunction with the troponins, as the components of actin-myosin contractile apparatus, is well-de®ned (Lin et al, 1997;Pittenger et al, 1994).…”

“…Restoration of TM1 expression abolishes the transformed phenotype induced by v-ki-ras oncogene (Prasad et al, 1993). Furthermore, enhanced expression of a closely related isoform, tropomyosin 2 (TM2), does not in¯uence the transformed phenotype of DT cells (Braverman et al, 1996).…”

“…The 3T3/src cells readily form colonies in agar, in anchorage independent fashion, as well as manifest morphological features concomitant with the transformed phenotype. In order to test whether the anti-oncogenic e ects of TM1 are speci®c to ras oncogene (Prasad et al, 1993), or whether the transformed phenotype induced by other functionally oncogenes could also be suppressed by TM1, we have restored the expression of TM1 in v-src transformed NIH3T3 cells.…”