Crystal Structure of CC3 (TIP30)

Abstract

CC3 (TIP30) is a protein with pro-apoptotic and antimetastatic properties. The tumor suppressor effect of CC3 has been suggested to result from inhibition of nuclear transport by binding to importin ␤s or by regulating transcription through interaction in a complex with co-activator independent of AF-2 function (CIA) and the c-myc gene. Previous biochemical studies indicated that CC3 has protein kinase activity, and a structural similarity to cAMP-dependent protein kinase catalytic subunit was proposed. By contrast, bioinformatics studies suggested a relationship of CC3 to the short chain dehydrogenase reductase family. To clarify details of the CC3 structural family and ligand binding properties, we have determined the crystal structure of CC3 at 1.7-Å resolution. CC3 has a short chain dehydrogenase reductase fold and binding specificity for NADPH, yet it is unlikely to be normally enzymatically active because it is monomeric. These structural results, in conjunction with data from earlier mutagenesis work on the nucleotide binding motif, suggest that NADPH binding is important for the biological activity of CC3, including interaction with importins and with the CIA/c-myc system. CC3 provides an example of the adaptation of a metabolic enzyme fold to include a regulatory role, as also seen in the case of the NADH-binding co-repressor CtBP.Apoptosis or programmed cell death plays an opposing role to cell division in the homeostatic regulation of animal cell populations. As well as being of significance in development, apoptosis has an important place in the pathogenesis of some disease states. Mutated and deleted pro-apoptotic genes can give rise to carcinogenesis and tumor growth (1). CC3 (also known as TIP30) is an anti-tumor protein that predisposes cells to apoptosis, thereby giving properties of metastatic suppression (2). CC3 is an evolutionarily conserved gene ubiquitously expressed as a 27-kDa protein in human tissues. CC3 RNA is absent in variant small cell lung carcinoma cell lines that are derived from tumors with highly aggressive metastatic behavior (2). Significantly, ectopic expression of CC3 in variant small cell lung carcinoma cells induces expression of a number of apoptosis-related genes including Bad and Siva (3) and also attenuates metastasis in vivo (2). Moreover, it has been shown that genetically engineered mice deficient in CC3 have a high incidence of liver cancer and that reduced expression of CC3 is also associated with 33% of human hepatocellular carcinomas (4).A number of studies have been aimed at defining the biochemical properties of CC3, as well as the interactions made with partner proteins that may relate to its pro-apoptotic effects. Recent results have shown that CC3 can bind directly to karyopherins of the importin ␤ family of nuclear transportins in a Ran-GTP-insensitive manner (5). For importin ␤2, the site of interaction with CC3 involves HEAT domain(s) in the Cterminal third of the protein. Binding of CC3 to importins inhibits the nuclear transport of proteins with eith...

An NADPH Sensor Protein (HSCARG) Down-regulates Nitric Oxide Synthesis by Association with Argininosuccinate Synthetase and Is Essential for Epithelial Cell Viability

“…In addition, a group of NAD/NADP-dependent proteins, including NmrA, CC3, and C-terminal binding protein, has been shown to regulate gene transcription factors by sensing the cellular level of NAD/NADP (30,(47)(48)(49)(50). Our observation that HSCARG enters the nucleus subsequent to a change in the intracellular redox state, together with our data that overexpression of HSCARG leads to NFB inhibition, 3 suggested that HSCARG may also target nuclear proteins, such as transcription factors, to further counter-balance the effects of the decreasing NADPH concentration.…”

Factors that retard remyelination in multiple sclerosis with a focus on TIP30: a novel therapeutic target

“…The crystal structure study of TIP30, however, failed to provide structural evidence for these bioactivities [96]. Rather, the study revealed that TIP30 can bind NADPH, which may influence its function, and may directly bind and block the cargo binding position of Importin b, an essential shuttle protein for nucleocytoplasmic transport [77], in a Ran-GTP-independent manner [96]. In support of this structural notion, biochemical studies [89,97] have shown that TIP30 indeed inhibits nucleocytoplasmic transport mediated by Importin b in a Ran-GTP-independent manner.…”

“…On the other hand, other bioinformatic studies have suggested that TIP30 is related to short-chain dehydrogenases/reductases (SDR) [94,95]. The crystal structure study of TIP30, however, failed to provide structural evidence for these bioactivities [96]. Rather, the study revealed that TIP30 can bind NADPH, which may influence its function, and may directly bind and block the cargo binding position of Importin b, an essential shuttle protein for nucleocytoplasmic transport [77], in a Ran-GTP-independent manner [96].…”

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“…Likewise, Gal3p, the transcriptional inducer in S. cerevisiae, demonstrates a ϳ90% sequence similarity to Gal1p, the bona fide galactokinase of the Leloir pathway that can, itself, function as a ligand sensor (24 -30). The molecular architecture of UDP-galactose 4-epimerase, another enzyme in galactose metabolism (31), has now been observed in NmrA, a negative transcriptional regulator in Aspergillus nidulans (32) and TIP30/CC3, a putative metastasis suppressor that promotes apoptosis (33). More than likely, other examples will be found where enzymatic scaffolds have been hijacked to serve in transcriptional regulation roles.…”