Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.108 (TAT)
 2,389 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Progression of cells through the G1 phase of the cell cycle requires cyclin D:Cdk4/6 and cyclin E:Cdk2 complexes; however, the duration and ordering of these complexes remain unclear. To address this, we synthesized a peptidyl mimetic of the Cdk4/6 inhibitor, p16INK4a that contained an NH2-terminal TAT protein transduction domain. Transduction of TAT-p16 wild-type peptides into cells resulted in the loss of active, hypophosphorylated pRb and elicited an early G1 cell cycle arrest, provided cyclin E:Cdk2 complexes were inactive. We conclude that cyclin D:Cdk4/6 activity is required for early G1 phase cell cycle progression up to, but not beyond, activation of cyclin E:Cdk2 complexes at the restriction point and is thus nonredundant with cyclin E:Cdk2 in late G1.
 ...
PMID:Transduced p16INK4a peptides inhibit hypophosphorylation of the retinoblastoma protein and cell cycle progression prior to activation of Cdk2 complexes in late G1. 1036 76

The retinoblastoma tumor suppressor protein (pRB) negatively regulates early-G(1) cell cycle progression, in part, by sequestering E2F transcription factors and repressing E2F-responsive genes. Although pRB is phosphorylated on up to 16 cyclin-dependent kinase (Cdk) sites by multiple G(1) cyclin-Cdk complexes, the active form(s) of pRB in vivo remains unknown. pRB is present as an unphosphorylated protein in G(0) quiescent cells and becomes hypophosphorylated (approximately 2 mol of PO(4) to 1 mol of pRB) in early G(1) and hyperphosphorylated (approximately 10 mol of PO(4) to 1 mol of pRB) in late G(1) phase. Here, we report that hypophosphorylated pRB, present in early G(1), represents the biologically active form of pRB in vivo that is assembled with E2Fs and E1A but that both unphosphorylated pRB in G(0) and hyperphosphorylated pRB in late G(1) fail to become assembled with E2Fs and E1A. Furthermore, using transducible dominant-negative TAT fusion proteins that differentially target cyclin D-Cdk4 or cyclin D-Cdk6 (cyclin D-Cdk4/6) and cyclin E-Cdk2 complexes, namely, TAT-p16 and TAT-dominant-negative Cdk2, respectively, we found that, in vivo, cyclin D-Cdk4/6 complexes hypophosphorylate pRB in early G(1) and that cyclin E-Cdk2 complexes inactivate pRB by hyperphosphorylation in late G(1). Moreover, we found that cycling human tumor cells expressing deregulated cyclin D-Cdk4/6 complexes, due to deletion of the p16(INK4a) gene, contained hypophosphorylated pRB that was bound to E2Fs in early G(1) and that E2F-responsive genes, including those for dihydrofolate reductase and cyclin E, were transcriptionally repressed. Thus, we conclude that, physiologically, pRB is differentially regulated by G(1) cyclin-Cdk complexes.
 ...
PMID:Differential regulation of retinoblastoma tumor suppressor protein by G(1) cyclin-dependent kinase complexes in vivo. 1141 52

Initiation of T-lymphocyte-mediated immune responses involves two cellular processes: entry into the cell cycle (G(0)-->G(1)) for clonal proliferation and coordinated changes in surface and secreted molecules that mediate effector functions. However, a point during G(0)-->G(1) beyond which T cells are committed to enter the cell cycle has not been defined. We define here a G(0)-->G(1) commitment point that occurs 3 to 5 h after CD3 and CD28 stimulation of human CD4 or CD8 T cells. Transition through this point requires cdk6/4-cyclin D, since inhibition with TAT-p16(INK4A) during the first 3 to 5 h prevents cell cycle entry and maintains both naive and memory T cells in G(0). Transition through the G(0)-->G(1) commitment point is also necessary for T cells to increase in size, i.e., to enter the cellular growth cycle. However, transition through this point is not required for the induction of effector functions. These can be initiated while cells are maintained in G(0) with TAT-p16(INK4A). We have termed this quiescent, activated state G(0(A)). Our data provide proof of the principle that entry of T cells into the cell cycle and cellular growth cycles are coupled at the G(0)-->G(1) commitment point but that these processes can be uncoupled from the early expression of molecules of effector functions.
 ...
PMID:Commitment point during G0-->G1 that controls entry into the cell cycle. 1264 Jan 20

Deregulation of the p16INK4a-cyclin D:cyclin-dependent kinases (cdk) 4/6-retinoblastoma (pRB) pathway is a common paradigm in the oncogenic transformation of human cells and suggests that this pathway functions linearly in malignant transformation. However, it is not understood why p16INK4a and cyclin D:cdk4/6 mutations are disproportionately more common than the rare genetic event of RB inactivation in human malignancies such as melanoma. To better understand how these complexes contribute to altered tissue homeostasis, we blocked cdk4/6 activation and acutely inactivated Rb by conditional mutagenesis during mouse hair follicle cycling. Inhibition of cdk4/6 in the skin by subcutaneous administration of a membrane-transducible TAT-p16INK4a protein completely blocked hair follicle growth and differentiation. In contrast, acute disruption of Rb in the skin of homozygous RbLoxP/LoxP mice via subcutaneous administration of TAT-Cre recombinase failed to affect hair growth. However, loss of Rb resulted in severe depigmentation of hair follicles. Further analysis of follicular melanocytes in vivo and in primary cell culture demonstrated that pRB plays a cell-autonomous role in melanocyte survival. Moreover, functional inactivation of all three Rb family members (Rb, p107, and p130) in primary melanocytes by treatment with a transducible TAT-E1A protein did not rescue the apoptotic phenotype. These findings suggest that deregulated cyclin D:cdk4/6 complexes and pRB perform nonoverlapping functions in vivo and provide a cellular mechanism that accounts for the low incidence of RB inactivation in cancers such as melanoma.
 ...
PMID:Distinct and nonoverlapping roles for pRB and cyclin D:cyclin-dependent kinases 4/6 activity in melanocyte survival. 1463 Sep 48

Peritoneal B-1a cells differ from splenic B-2 cells in the molecular mechanisms that control G(0)-S progression. In contrast to B-2 cells, cyclin D2 is up-regulated in a rapid and transient manner in phorbol ester (PMA)-stimulated B-1a cells, whereas cyclin D3 does not accumulate until late G(1) phase. This nonoverlapping expression of cyclins D2 and D3 suggests distinct functions for these proteins in B-1a cells. To investigate the contribution of cyclin D3 in the proliferation of B-1a cells, we transduced p16(INK4a) peptidyl mimetics (TAT-p16) into B-1a cells before cyclin D3 induction to specifically block cyclin D3-cyclin-dependent kinase 4/6 assembly. TAT-p16 inhibited DNA synthesis in B-1a cells stimulated by PMA, CD40L, or LPS as well as endogenous pRb phosphorylation by cyclin D-cyclin-dependent kinase 4/6. Unexpectedly, however, cyclin D3-deficient B-1a cells proliferated in a manner similar to wild-type B-1a cells following PMA or LPS stimulation. This was due, at least in part, to the compensatory sustained accumulation of cyclin D2 throughout G(0)-S progression. Taken together, experiments in which cyclin D3 was inhibited in real time demonstrate the key role this cyclin plays in normal B-1a cell mitogenesis, whereas experiments with cyclin D3-deficient B-1a cells show that cyclin D2 can compensate for cyclin D3 loss in mutant mice.
 ...
PMID:Disruption of cyclin D3 blocks proliferation of normal B-1a cells, but loss of cyclin D3 is compensated by cyclin D2 in cyclin D3-deficient mice. 1681 32

Endomitosis is a unique form of cell cycle used by megakaryocytes, in which the latter stages of mitosis are bypassed so that the cell can increase its DNA content and size. Although several transcription factors, including GATA-1 and RUNX-1, have been implicated in this process, the link between transcription factors and polyploidization remains undefined. Here we show that GATA-1-deficient megakaryocytes, which display reduced size and polyploidization, express nearly 10-fold less cyclin D1 and 10-fold increased levels of p16 compared with their wild-type counterparts. We further demonstrate that cyclin D1 is a direct GATA-1 target in megakaryocytes, but not erythroid cells. Restoration of cyclin D1 expression, when accompanied by ectopic overexpression of its partner Cdk4, resulted in a dramatic increase in megakaryocyte size and DNA content. However, terminal differentiation was not rescued. Of note, polyploidization was only modestly reduced in cyclin D1-deficient mice, likely due to compensation by elevated cyclin D3 expression. Finally, consistent with an additional defect conferred by increased levels of p16, inhibition of cyclin D-Cdk4 complexes with a TAT-p16 fusion peptide significantly blocked polyploidization of wild-type megakaryocytes. Together, these data show that GATA-1 controls growth and polyploidization by regulating cyclin D-Cdk4 kinase activity.
 ...
PMID:Cyclin D-Cdk4 is regulated by GATA-1 and required for megakaryocyte growth and polyploidization. 1731 55