The retinoblastoma family of proteins and their regulatory functions in the mammalian cell division cycle

Abstract

The retinoblastoma (RB) family of proteins are found in organisms as distantly related as humans, plants, and insects. These proteins play a key role in regulating advancement of the cell division cycle from the G1 to S-phases. This is achieved through negative regulation of two important positive regulators of cell cycle entry, E2F transcription factors and cyclin dependent kinases. In growth arrested cells transcriptional activity by E2Fs is repressed by RB proteins. Stimulation of cell cycle entry by growth factor signaling leads to activation of cyclin dependent kinases. They in turn phosphorylate and inactivate the RB family proteins, leading to E2F activation and additional cyclin dependent kinase activity. This propels the cell cycle irreversibly forward leading to DNA synthesis. This review will focus on the basic biochemistry and cell biology governing the regulation and activity of mammalian RB family proteins in cell cycle control.

Figure 1

Schematic representation of pRB, p107 and p130 open reading frames. (A) The central feature of RB-family proteins is the pocket domain. It was originally defined as the minimal domain necessary to bind to viral oncoproteins such as simian virus TAg through their LXCXE motif, and is denoted as the 'small pocket' in this diagram. The 'large pocket' is the minimal growth suppressing domain of RB-family proteins and it is capable of binding E2F transcription factors as well as viral proteins. (B) Comparison of open reading frame structures of each of the pocket proteins. Note the additional features found in the p107 and p130 proteins, the kinase inhibitory site, the cyclin binding site, and the insertion in the B-domain of the pocket. These provide the most obvious differences between pRB and its relatives p107 and p130.

Figure 2

Model of cell cycle entry control by pocket protein. Beginning in the top left corner, quiescent cells repress transcription of E2F targets genes largely through the actions of p130. As cells progress into G1, complexes containing p107 and a repressor E2F such as E2F4 begin to replace p130. Furthermore, complexes of pRB and activator E2Fs such as E2F3 also become more abundant. Chromatin remodeling factors (CRF) are recruited to these complexes and mediate alterations to the chromatin environment, preventing transcription of E2F responsive genes. As a result, transcription of E2F target genes remains low until entry into S-phase. At the transition to S-phase, cyclin/CDK complexes phosphorylate the pocket proteins, dissociating them from the E2F/DP duplexes and transcription of E2F target genes proceeds through S phase. As part of this transition, the repressive heterochromatin changes that were present in G1 are reversed by the recruitment of new enzymes by the E2Fs, histone acetyltransferases (HAT) are examples of this type of enzyme. Another important change at the start of S-phase is the export of p130 and 107 proteins from the nucleus. At this point pocket proteins are thought to be relatively functionless until they are dephosphorylated and reactivated at the end of mitosis so that they can regulate transcription again during the next G1 phase.

Figure 3

Expression levels of pocket proteins throughout the cell cycle. In G0, the most abundant pocket protein is p130. After cells are stimulated to enter the cell cycle expression of pRB and p107 are induced because they are E2F target genes themselves. At the same time these pocket proteins increase, the expression level of p130 begins to decline. In subsequent cell cycles pRB and p107 remain expressed at relatively constant levels, conversely, p130 is relatively inabundant under these growth conditions. These unique expression patterns offer clear, distinguishing characteristics of each pocket protein family member.

Figure 4

The interrelationship of cell cycle regulatory molecules in G1. Growth inhibitory and promoting signals impinge on the regulation of cyclin dependent kinase inhibitors (CKI). Growth promoting signals also directly lead to activation of cyclin dependent kinases (CDK) in G1. The cyclin dependent kinases serve to inactivate pocket proteins through phosphorylation leading to E2F transcriptional increases and cell cycle advancement. Cell cycle exit can be caused by activation of pocket proteins by phosphatases. In addition to blocking E2F transcription, recently activated pocket proteins also serve to negatively influence cyclin dependent kinase activity and positively influence CKI abundance. In this way, the regulatory molecules that control progression through G1 are extensively regulated by one another.