BTG2 (BTG Family Member-2) is endowed with antiproliferative activity. The expression of BTG2 in cycling cells induces accumulation of hypophosphorylated, growth-inhibitory forms of Rb (Retinoblastoma) and led to G1 arrest through impairment of DNA synthesis. Overexpression of CcnD1 (Cyclin-D1) counteracts G1 arrest. Rb is a nuclear phosphoprotein whose phosphorylation state oscillates regularly during the cell cycle. Its under-phosphorylated forms predominate in G0 and G1, while highly phosphorylated forms exist in S, G2 and M phases (Ref.1). The primary biological function of under-phosphorylated Rb is to inhibit progression toward S phase by controlling a checkpoint in late G1. In fact, under-phosphorylated Rb associates with members of the E2F family of transcription factors, impairing their activity and leading to a cell cycle block in G1. Conversely, the phosphorylation of Rb inactivates its growth suppression activity by freeing E2F molecules, thus enabling them to transactivate genes required for the progression of the cell into S phase and the remainder of the cell cycle. CDKs (Cyclin-Dependent Kinases) are the molecules responsible for Rb phosphorylation and its consequent inactivation (Ref.2).
BTG2 impairs G1-S progression, either by an Rb-dependent pathway through inhibition of CcnD1 transcription, or in an Rb-independent fashion by CcnE (Cyclin-E) down-regulation. BTG2 either inhibit synthesis of CcnE and CDK4 or accelerate degradation of CcnE through the Ubiquitination pathway with or without E2F. BTG2 also controls the G2 checkpoint. Furthermore, BTG2 interacts with CAF1 (Carbon Catabolite Repressor Protein (CCR4-Associative Factor-1), which influences cell cycle with the transcription factor HOXB9 (Homeobox-B9) and PRMT1 (Protein Arginine N-Methyl Transferase-1), that control transcription through Histone methylation (Ref.2). The molecular function of BTG2 is still unknown, but its ability to modulate CcnD1 transcription or to synergize with the transcription factor HOXB9 shows that it behaves as a transcriptional co-regulator. BTG1 and BTG2 bind and positively modulate the transcriptional activity of the HOXB9. BTG2 controls neuronal differerentiation by modulating the transcriptional activity of HOXB9. BTG3 has a colinearity of expression with HOXB9 certainly higher than BTG2. Formation of an active CcnD1-CDK4 complex depends on de novo synthesis of D1-protein. Inhibition of proliferation and down-regulation of CcnD1 levels by BTG2 events are significantly connected. Conversely, CcnD1 is able to rescue the BTG2-dependent G1 arrest, evidence pointing to CcnD1 as a target for the BTG2 inhibition of the cell cycle (Ref.1 & 3).
As for the mechanism by which BTG2 induces G1 arrest, relies on the inhibition of CcnD1 transcription, with consequent reduction of its protein levels. This impairs the ability of CcnD1-CDK4 to inactivate Rb through phosphorylation, the preliminary step that triggers the cell cycle entry in G1. BTG1 and BTG2 interact with PRMT1 and increase PRMT1 methyl-transferase activity. Box-C domain in BTG1 and BTG2 are responsible for the interaction. PRMT1 activity is essential for Growth Factor-induced cell differentiation, and blocking PRMT1 by Box-C domain of BTG1 or BTG2 induces apoptosis (Ref.3). The binding of PRMT1 to PP2A (Protein Phosphatase-2-Structural/RegulatorySubunit-A), which is implicated in several processes including DNA replication, extends to unexplored possibilities the likelihood of a control of cell cycle by BTG2 through modulation of protein methylation. BTG1 expression is high in G0-G1 and more generally in quiescent tissues, and is down-regulated as the cell enters the S phase, to return high in the following. Further association of BTG1 and BTG2 with CAF1 modulates transcriptional activity of CAF1. A CDK2-mediated phosphorylation on Ser159 of BTG1 is required for the binding of BTG1 to CAF1 and also for the antiproliferative activity of BTG1. CAF1 also binds to CDK4 and CDC2, showing that members of the BTG family regulate the cell cycle by modulating CDK
activities through their interaction with CAF1. CAF1 associates with CCRN4L (CCR4 Carbon Catabolite Repression 4-Like (S. cerevisiae)) in order to influence cell cycle (Ref.4).
This indicates that BTG1 acts as a growth arrest gene responsible for the maintenance of the quiescent state, unlike it appears for BTG2. BTG2 exerts an anti-apoptotic action in consequence of its ability to reduce CcnD1 levels, thus preventing Rb inactivation and re-entry into the cycle. Thus the character of a negative regulator of the cell cycle (not constitutively part of the division machinery) is conferred on BTG2 since it inhibits G1-S progression either by Rb dependent manner, through inhibition of CcnD1 transcription, or Rb-independent, by CcnE down-regulation. BTG2 is induced only when activated by specific cellular cues (which include a variety of stimuli, such as DNA damage or other types of cellular stress (e.g., Hypoxia), cellular differentiation and apoptosis) (Ref.3). The BTG proteins occur in organisms ranging from nematodes to humans, and a large body of evidence shows that BTG proteins are mediators of multiple antiproliferative activities along with cell growth control and differentiation. Although little is known about the biological functions of BTG proteins, it is likely that they constitute a family of functionally related genes that are involved in the control of the cell cycle. This protein family modulates transcription in response to multiple stimuli. In conclusion, BTG proteins are novel regulators of transcription that lack an intrinsic transactivation domain, but enhance the recruitment of coactivators (Ref.3 & 4).