Kiyoshi Ohtani

A third subunit, the gamma chain, of the human interleukin-2 receptor (IL-2R) was identified, and a complementary DNA clone encoding this member of the cytokine receptor family was isolated. The gamma chain is necessary for the formation of the high- and intermediate-affinity receptors, which consists of alpha beta gamma heterotrimers and beta gamma heterodimers, respectively. The IL-2R on murine fibroblastoid cells can be internalized after binding IL-2 only if the gamma chain is present; alpha and beta are insufficient for internalization. Thus, the gamma chain is an indispensable component of the functional IL-2R.

A variety of results point to the transcription factor E2F as a critical determinant of the G1/S-phase transition during the cell cycle in mammalian cells, serving to activate the transcription of a group of genes that encode proteins necessary for DNA replication. In addition, E2F activity appears to be directly regulated by the action of retinoblastoma protein (RB) and RB-related proteins and indirectly regulated through the action of G1 cyclins and associated kinases. We now show that the accumulation of G1 cyclins is regulated by E2F1. E2F binding sites are found in both the cyclin E and cyclin D1 promoters, both promoters are activated by E2F gene products, and at least for cyclin E, the E2F sites contribute to cell cycle-dependent control. Most important, the endogenous cyclin E gene is activated following expression of the E2F1 product encoded by a recombinant adenovirus vector. These results suggest the involvement of E2F1 and cyclin E in an autoregulatory loop that governs the accumulation of critical activities affecting the progression of cells through G1.

Both positive and negative signals govern the progression of cells from G1 into S phase, and a variety of data implicate the E2F transcription factor as a target for the action of one class of negative regulators, the Rb family of growth suppressors. We now find that the E2F1 gene, which encodes one of the components of E2F activity, is subject to autoregulatory control during progression from G0 to S phase and that this primarily reflects a negative control in G0 and early G1, a time when the majority of E2F activity exits as a complex with Rb family members. In addition, we find that deregulated expression of G1 cyclins in quiescent cells stimulates the E2F1 promoter and that this is augmented by coexpression of cyclin-dependent kinases in an E2F-dependent manner. We conclude that the E2F1 gene is a downstream target for G1 cyclin-dependent kinase activity, most likely as a consequence of phosphorylation of Rb family members, and that the autoregulation of E2F1 transcription may provide a sensitive switch for regulating the accumulation of E2F activity during the transition from G1 to S phase.

We examined the ability of the trans-acting factor p40tax of human T-cell leukemia virus type I (HTLV-I), which is thought to be a crucial molecule in T-cell transformation by HTLV-I, to activate expression of a set of endogenous cellular genes related to T-cell proliferation. For this purpose we established a subclone (JPX-9) of Jurkat cells that was stably transfected with an expression plasmid containing the p40tax gene, whose expression is definitely dependent on heavy-metal ions. Expression of the interleukin-2 receptor alpha chain in JPX-9 cells was induced in response to the induction of p40tax expression, as has been demonstrated by others in transient transfection experiments with Jurkat cells. In addition, we found that significant enhancement of expression of the nuclear proto-oncogene c-fos was closely associated with expression of p40tax. Continous enhancement in the level of c-fos mRNA was observed in the presence of p40tax. In contrast, mRNA levels of other nuclear proto-oncogenes (c-myc, c-myb, and c-jun) were not appreciably effected by the expression of p40tax. These results suggest that (i) in addition to the interleukin-2-interleukin-2 receptor system, cellular genes such as c-fos, which regulate normal T-cell growth, are also activated directly or indirectly by p40tax and (ii) p40tax-induced modulation of gene expression plays a crucial role in T-cell transformation by HTLV-I.

Mammalian cell growth is governed by regulatory activities that include the products of genes such as c-myc and ras that act early in G1, as well as the E2F family of transcription factors that accumulate later in G1 to regulate the expression of genes involved in DNA replication. Previous work has shown that the expression of the E2F1, E2F2, and E2F3 gene products is tightly regulated by cell growth. To further explore the mechanisms controlling accumulation of E2F activity, we have isolated genomic sequences flanking the 5' region of the E2F2 coding sequence. Various assays demonstrate promoter activity in this sequence that reproduces the normal control of E2F2 expression during a growth stimulation. Sequence comparison reveals the presence of a variety of known transcription factor binding sites, including E-box elements that are consensus Myc binding sites, as well as E2F binding sites. We demonstrate that the E-box elements, which we show can function as Myc-responsive sites, contribute in a positive fashion to promoter function. We also find that E2F-dependent negative regulation in quiescent cells plays a significant role in the cell growth-dependent control of the promoter, similar to the regulation of the E2F1 gene promoter.