Transforming Acidic Coiled-coil Research Articles

Characterization of Dysregulated Transcription in TP53-Mutated AML Subpopulations

Introduction: TP53 is a DNA binding protein that regulates transcription and exerts many functions critical to homeostasis. AML with mutated TP53 (TP53mt) is associated with dismal outcomes and resistance to all currently available therapies. Therefore, delineation of transcriptional dysregulation in TP53mt AML is critical for identification of vulnerabilities in this leukemia subtype. Leukemia cells are functionally heterogeneous, with many cells at diagnosis susceptible to chemotherapy, and a minority of resistant cells that persist despite treatment. Deeper understanding of all leukemia sub-populations is necessary for elucidating mechanisms of resistance. We hypothesized that sub-populations such as leukemia-stem cells (LSCs), and therapy-residual cells possess identifiable, targetable characteristics that drive resistance. We performed RNA-sequencing and compared differences in gene expression between these sub-populations. Methods: We collected 113 bone marrow samples from 39 patients (pts) with adverse risk AML, including 16 pts with TP53mt (41%), 14 pts with FLT3 mutations (FLT3mt), and 10 pts (26%) with other adverse risk. We performed RNA-sequencing on paired pre- and post-treatment, bulk and FACS-sorted samples. Mononuclear cells were flow-sorted for bulk (CD45dim) and LSCs (Lin-CD34+CD38-CD123+) from diagnostic samples. Post-treatment samples were sorted for bulk mononuclear cells and therapy-residual cells, determined based on patient-specific aberrant phenotype using multi-color flow cytometry analysis (Xu J et al., Clinics in laboratory medicine 2017). Samples with low-expression of housekeeping genes were excluded from the analysis. Differential expression was analyzed using DESeq2 and Gene Set Enrichment Analysis (GSEA) was performed using the HALLMARK gene set and considered statistically significant if q<0.1. Results: Regardless of genetic context, we found that LSCs were characterized by down regulation of immune-related pathways such (interferon-g response, allograft rejection, inflammatory response), and upregulation of oxidative phosphorylation and DNA repair among top dysregulated pathways when compared with bulk cells. Unlike FLT3mt, TP53mt LSCs had upregulation of MYC targets when compared with bulk cells. LSCs with TP53mt had a characteristic gene expression signature with a consistent upregulation of 11 genes (Figure 1). Among these genes, TACC3, a transforming acidic coiled-coil (TACC) family member could be a potentially druggable target given its role in protecting microtubule stability and centrosome integrity and recent identification of small molecule TACC3 inhibitors as an anticancer agent (Akbulut et al., Mol Cancer Ther 2020). When comparing differentially expressed pathways in bulk pre-treatment cells of non-responders (post-treatment bone marrow blasts >5%) vs responders (post-treatment bone marrow blasts >5%), we found upregulation of MYC targets, MTORC1 signaling, E2F targets and G2M checkpoint. We also found downregulation of immune-related pathways, namely allograft rejection and interferon-g response. This indicates the possibility of similar transcriptional mechanisms which lead to resistance and immune evasion in LSCs and in bulk populations resistant to therapy. Finally, we analyzed expression of immunotherapy targets with ongoing clinical trials in AML and compared expression of these targets to CD34+ cells from healthy donors as reference (Figure 2). We found differential expression of targets such as CD123, with high expression in bulk and leukemia subpopulations regardless of genotype, therefore implying the possibility of universally targeting AML with a relatively high therapeutic index. However, targets such as FLT3 or KIT had similar expression in AML compared with bone marrow cells from healthy donors therefore indicating the possibility of affecting normal hematopoiesis in therapies against these targets. Targets such as LAG3 and TIM3 were preferentially expressed in samples with FLT3mt. Conclusion: In summary, we show characteristic differences in gene expression associated with TP53mt LSCs and identify novel targets and potential mechanisms of resistance. We demonstrate pervasive downregulation of immune related pathways in LSCs or resistant leukemia. These results could impact future therapeutic strategies for these resistant leukemia subsets. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A Highly Potent TACC3 Inhibitor as a Novel Anticancer Drug Candidate.

TACC3, a transforming acidic coiled-coil (TACC) family member, is frequently upregulated in a broad spectrum of cancers, including breast cancer. It plays critical roles in protecting microtubule stability and centrosome integrity that is often dysregulated in cancers; therefore, making TACC3 a highly attractive therapeutic target. Here, we identified a new TACC3-targeting chemotype, BO-264, through the screening of in-house compound collection. Direct interaction between BO-264 and TACC3 was validated by using several biochemical methods, including drug affinity responsive target stability, cellular thermal shift assay, and isothermal titration calorimetry. BO-264 demonstrated superior antiproliferative activity to the two currently reported TACC3 inhibitors, especially in aggressive breast cancer subtypes, basal and HER2+, via spindle assembly checkpoint-dependent mitotic arrest, DNA damage, and apoptosis, while the cytotoxicity against normal breast cells was negligible. Furthermore, BO-264 significantly decreased centrosomal TACC3 during both mitosis and interphase. BO-264 displayed potent antiproliferative activity (∼90% have less than 1 μmol/L GI50 value) in the NCI-60 cell line panel compromising of nine different cancer types. Noteworthy, BO-264 significantly inhibited the growth of cells harboring FGFR3-TACC3 fusion, an oncogenic driver in diverse malignancies. Importantly, its oral administration significantly impaired tumor growth in immunocompromised and immunocompetent breast and colon cancer mouse models, and increased survival without any major toxicity. Finally, TACC3 expression has been identified as strong independent prognostic factor in breast cancer and strongly prognostic in several different cancers. Overall, we identified a novel and highly potent TACC3 inhibitor as a novel potential anticancer agent, inducing spindle abnormalities and mitotic cell death.

The evolving picture of the glioblastoma genome.

Survival rates for patients with glioblastoma multiforme (GBM), the most common malignant brain tumor, remain dismal despite decades of basic and clinical research. New treatment strategies are desperately needed. Consequently, the high-throughput genomic technologies developed to facilitate examination of cancer genomes have recently been focused on GBM, with goals of identifying novel molecular targets and developing a clinically-relevant biological classification of this deadly cancer. These genomic analyses have provided an unprecedented view of the complex genetic landscape of GBM, confirming the central role of both well-established and previously-unsuspected genes in the development of these tumors [1,2]. In addition, despite the plethora of genes found to be altered in GBMs, these data have allowed a semblance of order to be applied to these tumors through the identification of a small number of core pathways critical to GBM pathogenesis and the elucidation of discrete molecular subgroups of GBM [3]. The list of biologically-relevant GBM genes remains incomplete, however, due in large part to the logistical challenge of establishing the functional significance of the numerous genes found to be infrequently altered (between 1 and 5%) in these cancers. The detection of focal regions of recurrent copy number gain or loss (amplifications or homozygous deletions) can be a useful tool for the identification of candidate cancer genes. Using such a high-resolution genomic approach, Duncan et al. have identified two genes as potentially relevant to the emerging picture of the GBM genome, ERRFI1 (located at chr1p36) and TACC3 (located at chr4p16) [4]. Genomic alterations in these two regions have been reported in a number of human cancers, but the specific target genes of these copy number alterations are not known. The Digital Karotyping and SNP array data provided by Duncan et al. have provided initial evidence for two reasonable candidates. Importantly, given the significant challenge of distinguishing between altered genes which are drivers (i.e. genes that contribute to tumorigenesis) and those that are merely passengers (i.e. genes that accumulate alterations during tumor development but are not biologically relevant), preliminary studies of the potential functional significance of the candidate genes have been conducted. Recurrent focal deletions at 1p36, previously reported in numerous cancers including GBM, were found by Duncan et al. in their cohort of GBMs and validated using data from The Cancer Genome Atlas (TCGA) project. Two distinct minimal deleted regions were identified, with the most commonly-deleted region containing a single gene, ERRFI1. ERRFI1 expression was also found to be downregulated in ~34% of GBM samples analyzed, and functional studies of a cell line with a homozygous deletion of ERRFI1 revealed impaired wound-healing and a lower rate of trans-well migration after transfection with a vector containing ERRFI1, suggesting ERRFI1 as a potential tumor suppressor gene in GBMs. Similarly, focal gains at 4p16, most commonly involving TACC3, and overexpression of TACC3 were found in tumor samples, raising the possibility of TACC3 as a potential GBM oncogene. Both of these genes are biologically-plausible as GBM-related candidates. ERRFI1 acts as a negative regulator of EGFR and the ERbB family, while dysregulation of TACC (transforming acidic coiled coil) genes, components of the centrosome, has been implicated in cancer development. Significant work remains, however, in order to confirm the relevance of these genes for the pathogenesis of GBM and identify them as potential targets of molecular therapeutics. In particular, additional functional data firmly establishing the genes as “drivers” and demonstrating their specific biological role in GBM development are required. In addition, clarification of how these genomic data fit in with the evolving picture of the molecular classification of GBM will be illuminating: for example, are alterations of ERRFI1 specifically identified in the subgroup of GBMs with downstream evidence of EGFR activation that lack EGFR amplification or mutation [3]? Further research building on the foundation of this intriguing genomic data will surely add to our evolving picture of the glioblastoma genome. Hopefully it will also contribute to the improved care of our patients with these deadly cancers.

Absence of FGFR3–TACC3 rearrangement in hematological malignancies with numerical chromosomal alteration

FGFR–TACC, found in different tumor types, is characterized by the fusion of a member of fibroblast grown factor receptor (FGFR) tyrosine kinase (TK) family to a member of the transforming acidic coiled-coil (TACC) proteins. Because chromosome numerical alterations, hallmarks of FGFR–TACC fusions are present in many hematological disorders and there are no data on the prevalence, we studied a series of patients with acute myeloid leukemia and myelodysplastic syndrome who presented numerical alterations using cytogenetic traditional analysis. None of the analyzed samples showed FGFR3–TACC3 gene fusion, so screening for this mutation at diagnosis is not recommended.

The XMAP215 Ortholog Alp14 Promotes Microtubule Nucleation in Fission Yeast

The organization and number of microtubules (MTs) in a cell depend on the proper regulation of MT nucleation. Currently, the mechanism of nucleation is the most poorly understood aspect of MT dynamics. XMAP215/chTOG/Alp14/Stu2 proteins are MT polymerases that stimulate MT polymerization at MT plus ends by binding and releasing tubulin dimers. Although these proteins also localize to MT organizing centers and have nucleating activity invitro, it is not yet clear whether these proteins participate in MT nucleation invivo. Here, we demonstrate that in the fission yeast Schizosaccharomyces pombe, the XMAP215 ortholog Alp14 is critical for efficient MT nucleation invivo. In multiple assays, loss of Alp14 function led to reduced nucleation rate and numbers of interphase MT bundles. Conversely, activation of Alp14 led to increased nucleation frequency. Alp14 associated with Mto1 and γ-tubulin complex components, and artificially targeting Alp14 to the γ-tubulin ring complexes (γ-TuRCs) stimulated nucleation. In imaging individual nucleation events, we found that Alp14 transiently associated with a γ-tubulin particle shortly before the appearance of a new MT. The transforming acidic coiled-coil (TACC) ortholog Alp7 mediated the localization of Alp14 at nucleation sites but notplus ends, and was required for efficient nucleation but not for MT polymerization. Our findings provide the strongest evidence to date that Alp14 serves as a critical MT nucleation factor invivo. We suggest a model in which Alp14 associates with the γ-tubulin complex in an Alp7-dependent manner to facilitate the assembly or stabilization of the nascent MT.

Abstract 3019: The role of TACC3 in the progression from ductal carcinoma in situ to invasive breast cancer

Abstract Ductal carcinoma in situ (DCIS) is a non-invasive breast cancer with the potential to become invasive/metastatic. The development of DCIS to invasive breast cancer is an important aspect of breast tumor aggressiveness, however, little is known about the genetic and molecular alterations involved in this progression. Also, analysis of molecular markers and genetic signatures of DCIS and invasive breast cancer have failed to identify transition-specific biomarkers or pathways. Hence, identifying the molecular mechanisms by which DCIS progresses to invasive breast cancer is critical for the success of diagnosis, prognosis and therapy of breast cancer. Members of the transforming acidic coiled-coil (TACC) protein family are key players in the maintenance of centrosome integrity, the regulation of microtubule assembly, and spindle stability during mitosis. TACC3 has been shown to be involved in the regulation of centrosome-microtubule dynamic networks, cell growth and differentiation, and transcription/gene expression through interactions with other molecules. Mounting evidence indicates that high levels of TACC3 may contribute to breast cancer development. Here, we investigated the significance of TACC3 in the transition of DCIS to invasive breast cancer. First, we examined the expression of TACC3 in non-tumorigenic human mammary epithelial MCF10A cells and MCF10DCIS.COM cells, which are known to form DCIS-like lesions in mice and to progress to invasive cancer, and found that the expression of TACC3 is elevated in MCF10DCIS.COM cells compared to MCF10A cells. By analyzing tissue microarray data, we found that expression of TACC3 gradually increased from normal breast to DCIS, and further increased in invasive breast cancer. In addition, ectopic expression of TACC3 in MCF10DCIS.COM cells promoted cell proliferation, migratory and invasive capabilities as well as the epithelial-mesenchymal transition (EMT)-like phenotype. Moreover, MCF10DCIS.COM cells stably expressing TACC3 exhibited increased expression of stemness markers. Interestingly, a small molecule inhibitor of TACC3, KHS101 treatment in MCF10DCIS.COM cells significantly suppressed cell proliferation and the PI3K/AKT signaling pathway while having little effect on MCF10A cells. Our findings, therefore, suggest that TACC3 may play an important role in the progression from DCIS to invasive breast cancer and that targeting TACC3 could be a promising new therapeutic strategy for DCIS patients. Citation Format: Loredana Campo, Maya Mathew, Eun-Kyoung breuer, William Small. The role of TACC3 in the progression from ductal carcinoma in situ to invasive breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3019. doi:10.1158/1538-7445.AM2017-3019

TACC3 Is Important for Correct Progression of Meiosis in Bovine Oocytes.

Transforming acidic coiled-coil (TACC) proteins are key players during mitosis via stabilization of the spindle. The roles of TACCs during meiosis are however less clear. We used bovine oocytes to study the expression and function of TACC3 during meiosis. TACC3 mRNA was detected in bovine oocytes during meiosis using qRT-PCR, and while it was also expressed in cleavage stage embryos, its expression was down-regulated at the morula and blastocyst stages. Immunofluorescence was used to demonstrate that TACC3 co-localized with tubulin in the metaphase I and II spindles. However, TACC3 was not detected at anaphase or telophase of the first meiotic division. Aurora A, which is known to phosphorylate and activate TACC3 in mitotic cells, showed a similar pattern of gene expression to that of TACC3 in meiotic oocytes and preimplantation embryos. Aurora A protein was however only very transiently associated to the meiotic spindle. Pharmaceutical inhibition of Aurora A activity inhibited TACC3 phosphorylation but did not prevent TACC3 appearance in the spindle. Inhibiting Aurora A activity did however lead to abnormal meiotic spindle formation and impaired maturation of bovine oocytes. Similar results were obtained by knock-down of TACC3 expression using siRNA injection. These results suggest that TACC3 is important for stabilizing the meiotic spindle, but phosphorylation of TACC3 by Aurora A is not required for its recruitment to the meiotic spindle although phosphorylation of TACC3 by other kinases cannot be excluded.

Xenopus TACC1 is a microtubule plus-end tracking protein that can regulate microtubule dynamics during embryonic development.

Microtubule plus‐end dynamics are regulated by a family of proteins called plus‐end tracking proteins (+TIPs). We recently demonstrated that the transforming acidic coiled‐coil (TACC) domain family member, TACC3, can function as a +TIP to regulate microtubule dynamics in Xenopus laevis embryonic cells. Although it has been previously reported that TACC3 is the only TACC family member that exists in Xenopus, our examination of its genome determined that Xenopus, like all other vertebrates, contains three TACC family members. Here, we investigate the localization and function of Xenopus TACC1, the founding member of the TACC family. We demonstrate that it can act as a +TIP to regulate microtubule dynamics, and that the conserved C‐terminal TACC domain is required for its localization to plus‐ends. We also show that, in Xenopus embryonic mesenchymal cells, TACC1 and TACC3 are each required for maintaining normal microtubule growth speed but exhibit some functional redundancy in the regulation of microtubule growth lifetime. Given the conservation of TACC1 in Xenopus and other vertebrates, we propose that Xenopus laevis is a useful system to investigate unexplored cell biological functions of TACC1 and other TACC family members in the regulation of microtubule dynamics. © 2015 The Authors. Cytoskeleton, Published by Wiley Periodicals, Inc.

TACC3 Protein Regulates Microtubule Nucleation by Affecting γ-Tubulin Ring Complexes

Centrosome-mediated microtubule nucleation is essential for spindle assembly during mitosis. Although γ-tubulin complexes have primarily been implicated in the nucleation process, details of the underlying mechanisms remain poorly understood. Here, we demonstrated that a member of the human transforming acidic coiled-coil (TACC) protein family, TACC3, plays a critical role in microtubule nucleation at the centrosome. In mitotic cells, TACC3 knockdown substantially affected the assembly of microtubules in the astral region and impaired microtubule nucleation at the centrosomes. The TACC3 depletion-induced mitotic phenotype was rescued by expression of the TACC3 C terminus predominantly consisting of the TACC domain, suggesting that the TACC domain plays an important role in microtubule assembly. Consistently, experiments with the recombinant TACC domain of TACC3 demonstrated that this domain possesses intrinsic microtubule nucleating activity. Co-immunoprecipitation and sedimentation experiments revealed that TACC3 mediates interactions with proteins of both the γ-tubulin ring complex (γ-TuRC) and the γ-tubulin small complex (γ-TuSC). Interestingly, TACC3 depletion resulted in reduced levels of γ-TuRC and increased levels of γ-TuSC, indicating that the assembly of γ-TuRC from γ-TuSC requires TACC3. Detailed analyses suggested that TACC3 facilitates the association of γ-TuSC-specific proteins with the proteins known to be involved in the assembly of γ-TuRC. Consistent with such a role for TACC3, the suppression of TACC3 disrupted localization of γ-TuRC proteins to the centrosome. Our findings reveal that TACC3 is involved in the regulation of microtubule nucleation at the centrosome and functions in the stabilization of the γ-tubulin ring complex assembly.

XTACC3-XMAP215 association reveals an asymmetric interaction promoting microtubule elongation.

chTOG is a conserved microtubule polymerase that catalyses the addition of tubulin dimers to promote microtubule growth. chTOG interacts with TACC3, a member of the transforming acidic coiled-coil (TACC) family. Here we analyse their association using the Xenopus homologues, XTACC3 (TACC3) and XMAP215 (chTOG), dissecting the mechanism by which their interaction promotes microtubule elongation during spindle assembly. Using SAXS, we show that the TACC domain (TD) is an elongated structure that mediates the interaction with the C terminus of XMAP215. Our data suggest that one TD and two XMAP215 molecules associate to form a four-helix coiled-coil complex. A hybrid methods approach was used to define the precise regions of the TACC heptad repeat and the XMAP215 C terminus required for assembly and functioning of the complex. We show that XTACC3 can induce the recruitment of larger amounts of XMAP215 by increasing its local concentration, thereby promoting efficient microtubule elongation during mitosis.

TACC3 is a microtubule plus end-tracking protein that promotes axon elongation and also regulates microtubule plus end dynamics in multiple embryonic cell types.

Microtubule plus end dynamics are regulated by a conserved family of proteins called plus end-tracking proteins (+TIPs). It is unclear how various +TIPs interact with each other and with plus ends to control microtubule behavior. The centrosome-associated protein TACC3, a member of the transforming acidic coiled-coil (TACC) domain family, has been implicated in regulating several aspects of microtubule dynamics. However, TACC3 has not been shown to function as a +TIP in vertebrates. Here we show that TACC3 promotes axon outgrowth and regulates microtubule dynamics by increasing microtubule plus end velocities in vivo. We also demonstrate that TACC3 acts as a +TIP in multiple embryonic cell types and that this requires the conserved C-terminal TACC domain. Using high-resolution live-imaging data on tagged +TIPs, we show that TACC3 localizes to the extreme microtubule plus end, where it lies distal to the microtubule polymerization marker EB1 and directly overlaps with the microtubule polymerase XMAP215. TACC3 also plays a role in regulating XMAP215 stability and localizing XMAP215 to microtubule plus ends. Taken together, our results implicate TACC3 as a +TIP that functions with XMAP215 to regulate microtubule plus end dynamics.

Csi1p recruits alp7p/TACC to the spindle pole bodies for bipolar spindle formation.

Accurate chromosome segregation requires timely bipolar spindle formation during mitosis. The transforming acidic coiled-coil (TACC) family proteins and the ch-TOG family proteins are key players in bipolar spindle formation. They form a complex to stabilize spindle microtubules, mainly dependent on their localization to the centrosome (the spindle pole body [SPB] in yeast). The molecular mechanism underlying the targeting of the TACC-ch-TOG complex to the centrosome remains unclear. Here we show that the fission yeast Schizosaccharomyces pombe TACC orthologue alp7p is recruited to the SPB by csi1p. The csi1p-interacting region lies within the conserved TACC domain of alp7p, and the carboxyl-terminal domain of csi1p is responsible for interacting with alp7p. Compromised interaction between csi1p and alp7p impairs the localization of alp7p to the SPB during mitosis, thus delaying bipolar spindle formation and leading to anaphase B lagging chromosomes. Hence our study establishes that csi1p serves as a linking molecule tethering spindle-stabilizing factors to the SPB for promoting bipolar spindle assembly.

A small compound targeting TACC3 revealed its different spatiotemporal contributions for spindle assembly in cancer cells

The mitotic spindle is assembled by the coordinated action of centrosomes and kinetochore microtubules. An evolutionally conserved protein family, transforming acidic coiled-coil (TACC), has been shown to be involved in this process. In humans, TACC3 is aberrantly expressed in a variety of human cancers, but its biological significance remains to be elucidated. Here, using a novel compound targeting TACC3, spindlactone (SPL), we show that the perturbation of TACC3 selectively inhibited the nucleation of centrosome microtubules in ovarian cancer cells. In contrast to centrosome microtubules, the kinetochore microtubules were robustly assembled, forming ectopic spindle poles that resulted in multipolar spindles. Interestingly, the extensive inhibition of TACC3 partially suppressed the nucleation of kinetochore microtubules. These dose-dependent effects of SPL were consistent with the results observed by the depletion of TACC3 and its binding partner, colonic and hepatic tumor overexpressed gene protein (TOGp). Although these proteins both have roles in the assembly of centrosome and kinetochore microtubules, their contributions were spatiotemporally different. Notably, SPL did not affect spindle assembly in normal cells. Furthermore, the oral administration of SPL significantly suppressed tumor growth in vivo. The unique mechanism of action of SPL not only enables it to be used as a tool to dissect the molecular basis of spindle assembly but also to provide a rationale for the use of TACC3 as a molecular target for cancer treatment. This rationale offers an opportunity to develop new strategies for cancer chemotherapy that overcome the limitations of microtubule toxins and expand their scope and clinical efficacy.

Role of centrosomal adaptor proteins of the TACC family in the regulation of microtubule dynamics during mitotic cell division

During the mitotic division cycle, cells pass through an extensive microtubule rearrangement process where microtubules forming the mitotic spindle apparatus are dynamically instable. Several centrosomal- and microtubule-associated proteins are involved in the regulation of microtubule dynamics and stability during mitosis. Here, we focus on members of the transforming acidic coiled coil (TACC) family of centrosomal adaptor proteins, in particular TACC3, in which their subcellular localization at the mitotic spindle apparatus is controlled by Aurora-A kinase-mediated phosphorylation. At the effector level, several TACC-binding partners have been identified and characterized in greater detail, in particular, the microtubule polymerase XMAP215/ch-TOG/CKAP5 and clathrin heavy chain (CHC). We summarize the recent progress in the molecular understanding of these TACC3 protein complexes, which are crucial for proper mitotic spindle assembly and dynamics to prevent faulty cell division and aneuploidy. In this regard, the (patho)biological role of TACC3 in development and cancer will be discussed.

Repeats in Transforming Acidic Coiled-Coil (TACC) Genes

Transforming acidic coiled-coil proteins (TACC1, 2, and 3) are essential proteins associated with the assembly of spindle microtubules and maintenance of bipolarity. Dysregulation of TACCs is associated with tumorigenesis, but studies of microsatellite instability in TACC genes have not been extensive. Microsatellite or simple sequence repeat instability is known to cause many types of cancer. The present in silico analysis of SSRs in human TACC gene sequences shows the presence of mono- to hexa-nucleotide repeats, with the highest densities found for mono- and di-nucleotide repeats. Density of repeats is higher in introns than in exons. Some of the repeats are present in regulatory regions and retained introns. Human TACC genes show conservation of many repeat classes. Microsatellites in TACC genes could be valuable markers for monitoring numerical chromosomal aberrations and or cancer.

Transforming fusions induce aneuploidy

A recurrent oncogenic fusion of fibroblast growth factor receptor (FGFR) and transforming acidic coiled-coil (TACC) proteins in a subset of glioblastomas can directly interfere with cell division and induce aneuploidy, and this can be inhibited by FGFR kinase inhibitors.

Transforming fusions of FGFR and TACC genes in human glioblastoma.

The brain tumor glioblastoma multiforme (GBM) is among the most lethal forms of human cancer. Here, we report that a small subset of GBMs (3.1%; 3 of 97 tumors examined) harbors oncogenic chromosomal translocations that fuse in-frame the tyrosine kinase coding domains of fibroblast growth factor receptor (FGFR) genes (FGFR1 or FGFR3) to the transforming acidic coiled-coil (TACC) coding domains of TACC1 or TACC3, respectively. The FGFR-TACC fusion protein displays oncogenic activity when introduced into astrocytes or stereotactically transduced in the mouse brain. The fusion protein, which localizes to mitotic spindle poles, has constitutive kinase activity and induces mitotic and chromosomal segregation defects and triggers aneuploidy. Inhibition of FGFR kinase corrects the aneuploidy, and oral administration of an FGFR inhibitor prolongs survival of mice harboring intracranial FGFR3-TACC3-initiated glioma. FGFR-TACC fusions could potentially identify a subset of GBM patients who would benefit from targeted FGFR kinase inhibition.

The centrosomal protein TACC3 controls paclitaxel sensitivity by modulating a premature senescence program

Microtubule-interfering cancer drugs such as paclitaxel (PTX) often cause chemoresistance and severe side effects, including neurotoxicity. To explore potentially novel antineoplastic molecular targets, we investigated the cellular response of breast carcinoma cells to short hairpin(sh)RNA-mediated depletion of the centrosomal protein transforming acidic coiled coil (TACC) 3, an Aurora A kinase target expressed during mitosis. Unlike PTX, knockdown of TACC3 did not trigger a cell death response, but instead resulted in a progressive loss of the pro-apoptotic Bcl-2 protein Bim that links microtubule integrity to spindle poison-induced cell death. Interestingly, TACC3-depleted cells arrested in G₁ through a cellular senescence program characterized by the upregulation of nuclear p21(WAF), downregulation of the retinoblastoma protein and extracellular signal-regulated kinase 1/2, formation of HP1γ (phospho-Ser83)-positive senescence-associated heterochromatic foci and increased senescence-associated β-galactosidase activity. Remarkably, the onset of senescence following TACC3 knockdown was strongly accelerated in the presence of non-toxic PTX concentrations. Thus, we conclude that mitotic spindle stress is a major trigger of premature senescence and propose that the combined targeting of the centrosomal Aurora A-TACC3 axis together with drugs interfering with microtubule dynamics may efficiently improve the chemosensitivity of cancer cells.

The evolving picture of the glioblastoma genome.

Survival rates for patients with glioblastoma multiforme (GBM), the most common malignant brain tumor, remain dismal despite decades of basic and clinical research. New treatment strategies are desperately needed. Consequently, the high-throughput genomic technologies developed to facilitate examination of cancer genomes have recently been focused on GBM, with goals of identifying novel molecular targets and developing a clinically-relevant biological classification of this deadly cancer. These genomic analyses have provided an unprecedented view of the complex genetic landscape of GBM, confirming the central role of both well-established and previously-unsuspected genes in the development of these tumors [1,2]. In addition, despite the plethora of genes found to be altered in GBMs, these data have allowed a semblance of order to be applied to these tumors through the identification of a small number of core pathways critical to GBM pathogenesis and the elucidation of discrete molecular subgroups of GBM [3]. The list of biologically-relevant GBM genes remains incomplete, however, due in large part to the logistical challenge of establishing the functional significance of the numerous genes found to be infrequently altered (between 1 and 5%) in these cancers. The detection of focal regions of recurrent copy number gain or loss (amplifications or homozygous deletions) can be a useful tool for the identification of candidate cancer genes. Using such a high-resolution genomic approach, Duncan et al. have identified two genes as potentially relevant to the emerging picture of the GBM genome, ERRFI1 (located at chr1p36) and TACC3 (located at chr4p16) [4]. Genomic alterations in these two regions have been reported in a number of human cancers, but the specific target genes of these copy number alterations are not known. The Digital Karotyping and SNP array data provided by Duncan et al. have provided initial evidence for two reasonable candidates. Importantly, given the significant challenge of distinguishing between altered genes which are drivers (i.e. genes that contribute to tumorigenesis) and those that are merely passengers (i.e. genes that accumulate alterations during tumor development but are not biologically relevant), preliminary studies of the potential functional significance of the candidate genes have been conducted. Recurrent focal deletions at 1p36, previously reported in numerous cancers including GBM, were found by Duncan et al. in their cohort of GBMs and validated using data from The Cancer Genome Atlas (TCGA) project. Two distinct minimal deleted regions were identified, with the most commonly-deleted region containing a single gene, ERRFI1. ERRFI1 expression was also found to be downregulated in ~34% of GBM samples analyzed, and functional studies of a cell line with a homozygous deletion of ERRFI1 revealed impaired wound-healing and a lower rate of trans-well migration after transfection with a vector containing ERRFI1, suggesting ERRFI1 as a potential tumor suppressor gene in GBMs. Similarly, focal gains at 4p16, most commonly involving TACC3, and overexpression of TACC3 were found in tumor samples, raising the possibility of TACC3 as a potential GBM oncogene. Both of these genes are biologically-plausible as GBM-related candidates. ERRFI1 acts as a negative regulator of EGFR and the ERbB family, while dysregulation of TACC (transforming acidic coiled coil) genes, components of the centrosome, has been implicated in cancer development. Significant work remains, however, in order to confirm the relevance of these genes for the pathogenesis of GBM and identify them as potential targets of molecular therapeutics. In particular, additional functional data firmly establishing the genes as “drivers” and demonstrating their specific biological role in GBM development are required. In addition, clarification of how these genomic data fit in with the evolving picture of the molecular classification of GBM will be illuminating: for example, are alterations of ERRFI1 specifically identified in the subgroup of GBMs with downstream evidence of EGFR activation that lack EGFR amplification or mutation [3]? Further research building on the foundation of this intriguing genomic data will surely add to our evolving picture of the glioblastoma genome. Hopefully it will also contribute to the improved care of our patients with these deadly cancers.

TACC3-TSC2 maintains nuclear envelope structure and controls cell division

Studies of the role of tuberous sclerosis complex (TSC) proteins (TSC1/TSC2) in pathology have focused mainly on their capacity to regulate translation and cell growth, but their relationship with alterations of cellular structures and the cell cycle is not yet fully understood. The transforming acidic coiled-coil (TACC) domain-containing proteins are central players in structures and processes connected to the centrosome. Here, TACC3 interactome mapping identified TSC2 and 15 other physical interactors, including the evolutionary conserved interactions with ch-TOG/CKAP5 and FAM161B. TACC3 and TSC2 co-localize and co-purify with components of the nuclear envelope, and their deficiency causes morphological alterations of this structure. During cell division, TACC3 is necessary for the proper localization of phospho-Ser939 TSC2 at spindle poles and cytokinetic bridges. Accordingly, abscission alterations and increased frequency of binucleated cells were observed in Tacc3- and Tsc2-deficient cells relative to controls. In regulating cell division, TSC2 acts epistatically to TACC3 and, in addition to canonical TSC/mTOR signaling and cytokinetic associations, converges to the early mitotic checkpoint mediated by CHFR, consistently with nuclear envelope associations. Our findings link TACC3 to novel structural and cell division functions of TSC2, which may provide additional explanations for the clinical and pathological manifestations of lymphangioleiomyomatosis (LAM) disease and TSC syndrome, including the greater clinical severity of TSC2 mutations compared to TSC1 mutations.