E2F-4 and E2F-5, two members of the E2F family, are expressed in the early phases of the cell cycle.

Abstract

The E2F transcription factors play a role in regulating the expression of genes required for cell proliferation. Their activity appears to be regulated by association with the retinoblastoma protein (pRb) and the pRb-related proteins p107 and p130. In vivo, pRb is found in complex with a subset of E2F components-namely, E2F-1, E2F-2, and E2F-3. Here we describe the characterization of cDNAs encoding two unusual E2Fs, E2F-4 and E2F-5, each identified by the ability of their gene product to interact with p130 in a yeast two-hybrid system. E2F-4 and -5 share common sequences with E2F-1, E2F-2, and E2F-3 and, like these other E2Fs, the ability to heterodimerize with DP-1, thereby acquiring the ability to bind an E2F DNA recognition sequence with high affinity. However, in contrast to E2F-1, E2F-4 and E2F-5 fail to bind pRb in a two-hybrid assay. Moreover, they show a unique pattern of expression in synchronized human keratinocytes: E2F-4 and E2F-5 mRNA expression is maximal in mid-G1 phase before E2F-1 expression is detectable. These findings suggest that E2F-4 and E2F-5 may contribute to the regulation of early GI events including the Go/Gl transition. E2F/DP heterodimeric transcription factors are likely to be required for regulation of a large number of genes involved in cell proliferation (1, 2). An E2F consensus binding site has been demonstrated to be critical for the control of promoters activated at various different points in the cell cycle including the promoters of the c-myc (3, 4), DHFR (5), and cdc2 (6) genes. This wide spectrum of action may reflect the activities of several distinct E2F heterodimers, whose expression and function are regulated differentially, following distinct, cell cycle-specific schedules.A number of observations support this model. Cellular E2F activity is associated with several different protein species. Three distinct genes coding for E2Fs (7-11) and three for DPs (refs. 1 and 12; C. L. Wu and E. Harlow, personal communication) have already been identified. Moreover, the expression of the various E2Fs has been reported to be cell cycle dependent. For example, E2F-1 is expressed in the late G1 phase of the cell cycle (8, 13), clearly later than the induction of some E2F-responsive genes such as c-myc (3, 4). Finally, E2F activity appears to be directly and tightly regulated at several successive levels by the cell cycle machinery (14). For example, the E2F-1/DP-1 heterodimer appears to be inactivated through its binding to hypophosphorylated pRb in G, (15,16). Subsequently, phosphorylation of both pRb and E2F-1 (17) in late G1 results in the release of active E2F-1/ DP-1 transcription factors and in the transient expression of E2F-1-dependent genes. During the S and G2 phases that follow, the direct phosphorylation of E2F-1/DP-1 by cdk2/ cyclin A may then cause inactivation (14, 18). These processes may well explain the regulation of the three E2F subtypes (E2F-1, -2, and -3) that associate with pRb (11), but they do not address yet other aspects of E2F behavior. Thus, ...

Retinoblastoma family proteins as key targets of the small DNA virus oncoproteins

“…A-and B-domains are the most conserved among the three RB proteins and are involved in common functional characteristics (Paggi et al, 1996;Mulligan and Jacks, 1998), the most relevant one being the ability to control the cell cycle by negative modulation of the transition between the G1 and S phases (Goodrich et al, 1991;Zhu et al, 1993;Claudio et al, 1994;Starostik et al, 1996). To perform this task, these 'pocket proteins' utilize mechanisms mostly related to inactivation of transcription factors (Kouzarides, 1995), such as those of the E2F family (Cao et al, 1992;Helin et al, 1992Helin et al, , 1993Shirodkar et al, 1992;Beijersbergen et al, 1994;Hijmans et al, 1995;Sardet et al, 1995;Hurford et al, 1997), that promote the cell entrance into the S phase (Ewen, 1994;Weinberg, 1995;Paggi et al, 1996;Mulligan and Jacks, 1998). Indeed, in addition to the cell cycle, the RB family proteins regulate a wide spectrum of complex biological phenomena, as differentiation, embryonic development, apoptosis and senescence ( (Riley et al, 1994;Sidle et al, 1996;Herwig and Strauss, 1997;Stiegler et al, 1998;Thomas et al, 2003;Liu et al, 2004;Kapic et al, 2006) and references therein).…”

Identification of a p130 domain mediating interactions with cyclin A/cdk 2 and cyclin E/cdk 2 complexes

“…Particularly noteworthy among these are the interactions with the E2F/DP family of transcription factors (Cao et al, 1992;Devoto et al, 1992;Helin et al, 1992Helin et al, , 1993Kaelin et al, 1992;Shan et al, 1992;Shirodkar et al, 1992;Cobrinik et al, 1993;Ivey-Hoyle et al, 1993;Lees et al, 1993;Bandara et al, 1994;Bejersbergen et al, 1994;Ginsberg et al, 1994;Buck et al, 1995;Hijmans et al, 1995;Ormondroyd et al, 1995;Sardet et al, 1995;Wu et al, 1995;Zhang and Chellappan, 1995). The E2F transcription factors are thought to control expression of a number of genes involved in cell cycle progression (reviewed in Cobrinik, 1996).…”

K-ras modulates the cell cycle via both positive and negative regulatory pathways

“…An important element of this cyclin regulated cell cycle is the rapid induction of cell cycle phase speci®c cyclin expression and facile destruction of these cyclins as cells progress through the next phase of the cell cycle. In mammalian cells, the D-type cyclins (reviewed in Sherr, 1995), in association with the cyclin-dependent kinases cdk4 or cdk6, regulate the progression of the G 1 phase of the cell cycle mainly by participating in the phosphorylation of the retinoblastoma gene product pRb (reviewed in Weinberg, 1995). As a result of phosphorylation of pRb, free E2F, of which ®ve family members have been identi®ed to date (Sardet et al, 1995;, is released from complexes containing E2F and the hypophosphorylated form of pRb.…”

“…In mammalian cells, the D-type cyclins (reviewed in Sherr, 1995), in association with the cyclin-dependent kinases cdk4 or cdk6, regulate the progression of the G 1 phase of the cell cycle mainly by participating in the phosphorylation of the retinoblastoma gene product pRb (reviewed in Weinberg, 1995). As a result of phosphorylation of pRb, free E2F, of which ®ve family members have been identi®ed to date (Sardet et al, 1995;, is released from complexes containing E2F and the hypophosphorylated form of pRb. The`free' E2F, which is present as a heterodimer with its binding partner DP-1 or DP-2 (Wu et al, 1995), is the active transcription factor that promotes the transcription of E2F target genes such as dihydrofolate reductase (DHFR) (Blake and Azizkhan, 1989), thymidylate synthase (TS) (Johnson, 1994) and other genes necessary for entry into and progression through the S phase of the cell cycle (Adams and Kaelin, 1995;DeGregori et al, 1995).…”