Centrin Research Articles

Astrocytic centrin-2 expression in entorhinal cortex correlates with Alzheimer's disease severity.

Astrogliosis is a condition shared by acute and chronic neurological diseases and includes morphological, proteomic, and functional rearrangements of astroglia. In Alzheimer's disease (AD), reactive astrocytes frame amyloid deposits and exhibit structural changes associated with the overexpression of specific proteins, mostly belonging to intermediate filaments. At a functional level, amyloid beta triggers dysfunctional calcium signaling in astrocytes, which contributes to the maintenance of chronic neuroinflammation. Therefore, the identification of intracellular players that participate in astrocyte calcium signaling can help unveil the mechanisms underlying astrocyte reactivity and loss of function in AD. We have recently identified the calcium-binding protein centrin-2 (CETN2) as a novel astrocyte marker in the human brain and, in order to determine whether astrocytic CETN2 expression and distribution could be affected by neurodegenerative conditions, we examined its pattern in control and sporadic AD patients. By immunoblot, immunohistochemistry, and targeted-mass spectrometry, we report a positive correlation between entorhinal CETN2 immunoreactivity and neurocognitive impairment, along with the abundance of amyloid depositions and neurofibrillary tangles, thus highlighting a linear relationship between CETN2 expression and AD progression. CETN2-positive astrocytes were dispersed in the entorhinal cortex with a clustered pattern and colocalized with reactive glia markers STAT3, NFATc3, and YKL-40, indicating a human-specific role in AD-induced astrogliosis. Collectively, our data provide the first evidence that CETN2 is part of the astrocytic calcium toolkit undergoing rearrangements in AD and adds CETN2 to the list of proteins that could play a role in disease evolution.

Primary Ciliary Dyskinesia Associated Disease-Causing Variants in CCDC39 and CCDC40 Cause Axonemal Absence of Inner Dynein Arm Heavy Chains DNAH1, DNAH6, and DNAH7.

Disease-causing bi-allelic DNA variants in CCDC39 and CCDC40 are frequent causes of the hereditary disorder of primary ciliary dyskinesia (PCD). The encoded proteins form a molecular ruler complex, crucial for maintaining the 96 nm repeat units along the ciliary axonemes. Defects of those proteins cause a stiff, rapid, and flickery ciliary beating pattern, recurrent respiratory infections, axonemal disorganization, and abnormal assembly of GAS8, CCDC39, and DNALI1. We performed molecular characterization of the defects in the 96 nm axonemal ruler due to disease-causing variants in CCDC39 and CCDC40 and analyzed the effect on additional axonemal components. We identified a cohort of 51 individuals with disease-causing variants in CCDC39 and CCDC40 via next-generation sequencing techniques and demonstrated that the IDA heavy chains DNAH1, DNAH6, and DNAH7 are conspicuously absent within the respiratory ciliary axonemes by immunofluorescence analyses. Hence, we show for the first time that the centrin2 (CETN2) containing IDAs are also affected. These findings underscore the crucial role of CCDC39 and CCDC40 in the assembly and function of IDAs in human respiratory cilia. Thus, our data improve the diagnostics of axonemal ruler defects by further characterizing the associated molecular IDA defects.

The Chlamydia trachomatis type III-secreted effector protein CteG induces centrosome amplification through interactions with centrin-2

The centrosome is the main microtubule organizing center of the cell and is crucial for mitotic spindle assembly, chromosome segregation, and cell division. Centrosome duplication is tightly controlled, yet several pathogens, most notably oncogenic viruses, perturb this process leading to increased centrosome numbers. Infection by the obligate intracellular bacterium Chlamydia trachomatis (C.t.) correlates with blocked cytokinesis, supernumerary centrosomes, and multipolar spindles; however, the mechanisms behind how C.t. induces these cellular abnormalities remain largely unknown. Here we show that the secreted effector protein, CteG, binds to centrin-2 (CETN2), a key structural component of centrosomes and regulator of centriole duplication. Our data indicate that both CteG and CETN2 are necessary for infection-induced centrosome amplification, in a manner that requires the C-terminus of CteG. Strikingly, CteG is important for invivo infection and growth in primary cervical cells but is dispensable for growth in immortalized cells, highlighting the importance of this effector protein to chlamydial infection. These findings begin to provide mechanistic insight into how C.t. induces cellular abnormalities during infection, but also indicate that obligate intracellular bacteria may contribute to cellular transformation events. Centrosome amplification mediated by CteG-CETN2 interactions may explain why chlamydial infection leads to an increased risk of cervical or ovarian cancer.

High expression of CETN2 is associated with platinum resistance and poor prognosis in epithelial ovarian cancer.

The poor prognosis of ovarian cancer is largely due to platinum resistance. It has been demonstrated that nucleotide excision repair (NER) involving centrin-2(CETN2) is connected to platinum resistance in ovarian cancer. The molecular mechanism of CETN2 in ovarian cancer and the mechanism affecting the outcome of chemotherapy are unknown. The protein-protein interaction (PPI) network was mapped after obtaining the interacting proteins of CETN2, and the interacting genes were subjected to enrichment analysis. To examine the relationship between CETN2 and platinum resistance, gene microarray data and clinical data related to platinum resistance in ovarian cancer were downloaded. The possible signaling pathway of CETN2 was investigated by Gene set enrichment analysis (GSEA). Immune infiltration analysis was performed. Immunohistochemistry (IHC) and quantitative real-time PCR (QRT-PCR) were used to examine the expression of CETN2 in clinical samples in relation to the effectiveness of chemotherapy. The capacity of CETN2 to predict chemotherapy results was proven by receiver operating characteristic (ROC) curves after the construction of two prediction models, the logistic regression model and the decision tree model. The impact of CETN2 on prognosis was examined using the Kaplan-Meier technique. CETN2 was associated with NER, oxidative phosphorylation (OXPHOS) and cell cycle pathways in ovarian cancer drug-resistant samples. In clinical samples, CETN2 showed its possible correlation with immune infiltration. The protein expression level of CETN2 was significantly higher in platinum-resistant patients than that in platinum-sensitive patients, and the expression level had some predictive value for chemotherapy outcome, and high CETN2 protein expression was associated with poorer progression-free survival. CETN2 protein had a significant effect on ovarian cancer platinum sensitivity and prognosis, which may be related to the activation of NER, OXPHOS and cell cycle pathways upon CETN2 upregulation. Further research is necessary to determine the therapeutic application value of CETN2, which may be a new biomarker of chemoresponsiveness.

Centrin 2: A Novel Marker of Mature and Neoplastic Human Astrocytes.

As microtubule-organizing centers (MTOCs), centrosomes play a pivotal role in cell division, neurodevelopment and neuronal maturation. Among centrosomal proteins, centrin-2 (CETN2) also contributes to DNA repair mechanisms which are fundamental to prevent genomic instability during neural stem cell pool expansion. Nevertheless, the expression profile of CETN2 in human neural stem cells and their progeny is currently unknown. To address this question, we interrogated a platform of human neuroepithelial stem (NES) cells derived from post mortem developing brain or established from pluripotent cells and demonstrated that while CETN2 retains its centrosomal location in proliferating NES cells, its expression pattern changes upon differentiation. In particular, we found that CETN2 is selectively expressed in mature astrocytes with a broad cytoplasmic distribution. We then extended our findings on human autoptic nervous tissue samples. We investigated CETN2 distribution in diverse anatomical areas along the rostro-caudal neuraxis and pointed out a peculiar topography of CETN2-labeled astrocytes in humans which was not appreciable in murine tissues, where CETN2 was mostly confined to ependymal cells. As a prototypical condition with glial overproliferation, we also explored CETN2 expression in glioblastoma multiforme (GBM), reporting a focal concentration of CETN2 in neoplastic astrocytes. This study expands CETN2 localization beyond centrosomes and reveals a unique expression pattern that makes it eligible as a novel astrocytic molecular marker, thus opening new roads to glial biology and human neural conditions.

Transcriptional profile of ovine oocytes matured under lipopolysaccharide treatment in vitro

Lipopolysaccharide (LPS) derived from gram negative bacteria cell wall is known to cause ruminal acidosis and/or infectious diseases such as metritis and mastitis which has a significant negative impact on the reproductive performance. This study aimed to investigate the effect of LPS on oocyte maturation and subsequent development in vitro. Ovine cumulus oocyte complexes (COCs) were matured in a medium supplemented with 0 (control), 0.01, 0.1, 1 and 10 μg/mL LPS. Nuclear maturation, cleavage and blastocyst rate, mitochondrial membrane potential (ΔΨm), intracellular reactive oxygen species (ROS) content and changes to the transcript abundance were evaluated. In case of the maturation rate, the percentage of oocytes reaching the MII stage was lower following exposure to 10 μg/mL LPS in comparison to the control group (P < 0.05). Moreover, the blastocyst rate decreased in case of 1 and 10 μg/mL LPS when compared to the control group (P < 0.05). ROS overproduction accompanied by a decreased ΔΨm were recorded in LPS treated oocytes in comparison to the control group (P < 0.05). The 3′ tag digital gene expression profiling method revealed that 7887 genes were expressed while only seven genes exhibited changes in the transcript abundance following exposure to LPS. Tripartite motif containing 25 (TRIM25), Tripartite motif containing 26 (TRIM26), Zona Pellucida glycoprotein 3 (ZP3), Family with sequence similarity 50-member A (FAM50A), Glyoxalate and hydroxy pyruvate reductase (GRHPR), NADH ubiquinase oxireductase subunit A8 (NDUFA8) were down-regulated (P < 0.05), while only Centrin 3 (CETN3) was up-regulated (P < 0.05). Our findings show that LPS has undesirable effects on the maturation competence of ovine oocytes and subsequent embryo development. In addition, the transcriptomic profiling results may shed more light on the molecular mechanisms of LPS-induced infertility in ruminants.

Evaluation of Novel Antibodies to Centrin-1 for Radioimmunotherapy of Pancreatic Cancer

Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer death in the US with a very low survival rate. Unlike other solid malignancies, a biopsy of the pancreas is very invasive and recommend only with a mass suspected to be PDAC. Centrin1 (CETN1), a cancer/testis antigens (CATs), has been showed a 25-fold upregulation in 50% of the tumors from pancreatic cancer patients. Since testes are an immunoprivileged site, CETN1 could be a perfect target of radioimmunotherapy as the side effects of the treatment would be minimal. In this study, we developed novel antibodies (69-11 and 76-6) that are highly specific to CETN1, compared to its compensatory protein CENT2, which is widely expressed in all eukaryotic cells. 69-11 and 76-6 are either labeled with 213Bi, an alpha emitter, or with 177Lu, a beta emitter, for the treatment study. The radiolabeled antibodies were administered to PDAC xenografts-bearing nude mice. The localization of the radiolabeled antibodies in the tumors and normal organs was determined with micro SPECT/CT imaging. The tumors were monitored for 50 days. The toxicity assessment included weekly blood chemistry and kidney and liver functions assessment when PDAC-bearing mice were sacrificed at the end of the study. Labeling with 213Bi converted CETN1-specific antibodies into a very effective radioimmunotherapy reagent with tumor growth significantly (P=0.01) slowed down by either 100 or 200 μCi single injection. Importantly, the effect of the antibodies on the tumors was CETN1-specific, as 200 μCi control IgG had no effect on the tumor growth. In spite of impressive localization in the tumor demonstrated during the imaging experiments, 177Lu-labeled antibody was not very effective in slowing down tumor growth with no difference from 177Lu-IgG control (P=0.06) and was several folds less effective than 213Bi-labeled antibody. Both 213Bi and 177Lu groups showed only transient hematologic toxicity and absence of liver and kidney toxicity attesting to the very high safety margin of targeting CETN1 with radioimmunotherapy. In conclusion, the novel antibodies have the ability to detect CETN1 in vivo and in vitro, are highly efficacious and safe for treatment of PDAC, and warrant further work on developing them into clinical agents for diagnosis and therapy of PDAC.

Proteomic alterations underlie an association with teratozoospermia in obese mice sperm

BackgroundObesity is a worldwide crisis impairing human health. In this condition, declines in sperm quality stem from reductions in sperm concentration, motility and increase in sperm deformity. The mechanism underlying these alterations remains largely unknown. This study, determined if obesity-associated proteomic expression patterns in mice sperm parallel those in spermatozoa obtained from obese humans.MethodsAn obese mouse model was established via feeding a high-fat diet (HFD). Histological analysis identified testicular morphology and a computer assisted semen analyzer (CASA) evaluated sperm parameters. Proteome analysis was performed using a label-free quantitative LC-MS/MS system. Western blot, immunohistochemical and immunofluorescent analyses characterized protein expression levels and localization in testis, sperm and clinical samples.ResultsBodyweight gains on the HFD induced hepatic steatosis. Declines in sperm motility accompanied sperm deformity development. Differential proteomic analysis identified reduced cytoskeletal proteins, centrosome and spindle pole associated protein 1 (CSPP1) and Centrin 1 (CETN1), in sperm from obese mice. In normal weight mice, both CSPP1 and CETN1 were localized in the spermatocytes and spermatids. Their expression was appreciable in the post-acrosomal region parallel to the microtubule tracks of the manchette structure in spermatids, which affects spermatid head shaping and morphological maintenance. Moreover, CSPP1 was localized in the head–tail coupling apparatus of the mature sperm, while CETN1 expression was delimited to the post-acrosomal region within the sperm head. Importantly, sperm CSPP1 and CETN1 abundance in both the overweight and obese males decreased in comparison with that in normal weight men.ConclusionThese findings show that regionally distinct expression and localization of CETN1 and CSPP1 is strongly related to spermiogenesis and sperm morphology maintaining. Obesity is associated with declines in the CETN1 and CSPP1 abundance and compromise of both sperm morphology in mice and relevant clinical samples. This parallelism between altered protein expression in mice and humans suggests that these effects may contribute to poor sperm quality including increased deformity.

Evaluation of novel highly specific antibodies to cancer testis antigen Centrin-1 for radioimmunoimaging and radioimmunotherapy of pancreatic cancer.

BackgroundPancreatic ductal adenocarcinoma (PDAC) accounts for >90% of pancreatic malignancies, and has median survival of <6 months. There is an urgent need for diagnostic and therapeutic options for PDAC. Centrin1 (CETN1) is a novel member of Cancer/Testis Antigens, with a 25‐fold increase of CETN1 gene expression in PDX from PDAC patients. The absence of selective anti‐CETN1 antibodies is hampering CETN1 use for diagnosis and therapy. Here we report the generation of highly specific for CETN1 antibodies and their evaluation for radioimmunoimaging and radioimmunotherapy (RIT) of experimental PDAC.MethodsThe antibodies to CETN1 were generated via mice immunization with immunogenic peptide distinguishing CETN1 from CETN2. Patient tumor microarrays were used to evaluate the binding of the immune serum to PDAC versus normal pancreas. The antibodies were tested for their preferential binding to CETN1 over CETN2 by ELISA. Mice bearing PDAC MiaPaCa2 xenografts were imaged with microSPECT/CT and treated with 213Bi‐ and 177Lu‐labeled antibodies to CETN1.ResultsImmune serum bind to 50% PDAC cases on patient tumor microarrays with no specific binding to normal pancreas. Antibodies demonstrated preferential binding to CETN1 versus CETN2. Antibody 69‐11 localized to PDAC xenografts in mice in vivo and ex vivo. RIT of PDAC xenografts with 213Bi‐labeled antibodies was effective, safe, and CETN1‐specific.ConclusionsThe results demonstrate the ability of these novel antibodies to detect CETN1 both in vitro and in vivo; as well, the RIT treatment of experimental PDAC when radiolabeled with 213Bi is highly efficient and safe. Further evaluation of these novel reagents for diagnosis and treatment of PDAC is warranted.

The roles of Centrin 2 and Dynein Light Chain 8a in apical secretory organelles discharge of Toxoplasma gondii.

To efficiently enter host cells, apicomplexan parasites such as Toxoplasma gondii rely on an apical complex composed of tubulin-based structures as well as two sets of secretory organelles named micronemes and rhoptries. The trafficking and docking of these organelles to the apical pole of the parasite is crucial for the discharge of their contents. Here, we describe two proteins typically associated with microtubules, Centrin 2 (CEN2) and Dynein Light Chain 8a (DLC8a), that are required for efficient host cell invasion. CEN2 localizes to four different compartments, and remarkably, conditional depletion of the protein occurs in stepwise manner, sequentially depleting the protein pools from each location. This phenomenon allowed us to discern the essential function of the apical pool of CEN2 for microneme secretion, motility, invasion and egress. DLC8a localizes to the conoid, and its depletion also perturbs microneme exocytosis in addition to the apical docking of the rhoptry organelles, causing a severe defect in host cell invasion. Phenotypic characterization of CEN2 and DLC8a indicates that while both proteins participate in microneme secretion, they likely act at different steps along the cascade of events leading to organelle exocytosis.

Deletion of both centrin 2 (CETN2) and CETN3 destabilizes the distal connecting cilium of mouse photoreceptors

Centrins (CETN1-4) are ubiquitous and conserved EF-hand-family Ca2+-binding proteins associated with the centrosome, basal body, and transition zone. Deletion of CETN1 or CETN2 in mice causes male infertility or dysosmia, respectively, without affecting photoreceptor function. However, it remains unclear to what extent centrins are redundant with each other in photoreceptors. Here, to explore centrin redundancy, we generated Cetn3GT/GT single-knockout and Cetn2-/-;Cetn3GT/GT double-knockout mice. Whereas the Cetn3 deletion alone did not affect photoreceptor function, simultaneous ablation of Cetn2 and Cetn3 resulted in attenuated scotopic and photopic electroretinography (ERG) responses in mice at 3 months of age, with nearly complete retina degeneration at 1 year. Removal of CETN2 and CETN3 activity from the lumen of the connecting cilium (CC) destabilized the photoreceptor axoneme and reduced the CC length as early as postnatal day 22 (P22). In Cetn2-/-;Cetn3GT/GT double-knockout mice, spermatogenesis-associated 7 (SPATA7), a key organizer of the photoreceptor-specific distal CC, was depleted gradually, and CETN1 was condensed to the mid-segment of the CC. Ultrastructural analysis revealed that in this double knockout, the axoneme of the CC expanded radially at the distal end, with vertically misaligned outer segment discs and membrane whorls. These observations suggest that CETN2 and CETN3 cooperate in stabilizing the CC/axoneme structure.

ACTG1 and TLR3 are biomarkers for alcohol-associated hepatocellular carcinoma.

Alcohol consumption is a risk factor for the development of hepatocellular carcinoma (HCC); however, the association between alcohol and HCC remains unknown. The present study aimed to identify key genes related to alcohol-associated HCC to improve the current understanding of the pathology of this disease. Alcohol-associated and non-alcohol-associated HCC samples in the GSE50579 dataset of the Gene Omnibus Database were analyzed to investigate altered gene expression. Integrated bioinformatics methods were employed to clarify the biological functions of the differentially expressed genes (DEGs), including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein-protein interactions (PPIs). The present study reported that candidate biomarker micro (mi)RNAs via TargetScan Human 7.1. DEGs and their associated miRNAs (according to bioinformatics analysis) were validated using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Additionally, 284 EGs from the GSE50579 dataset were revealed. In GO term analysis, DEGs were closely associated with the ‘regulation of nucleic acid metabolism’. KEGG pathway analysis indicated that the DEGs were tightly engaged in the ‘VEGF and VEGF receptor signaling network’, ‘proteoglycan syndecan-mediated signaling events’, ‘erbB receptor signaling’ and ‘β1 integrin cell surface interactions’. According to the results of PPI and heat map analysis, the main hub genes were centrin 3 (CETN3), Toll-like receptor 3 (TLR3), receptor tyrosine-protein kinase (ERBB4), heat shock protein family member 8, actin γ1 (ACTG1) and α-smooth muscle actin. it was demonstrated that the ACTG1, TLR3, miR-6819-3p and miRΝΑ (miR)-6877-3P had undefined associations. Furthermore, RT-qPCR analysis revealed that miR-6819-3p and miR-6877-3P may enhance the expression levels of ACTG1 and inhibit the expression levels of TLR3 in alcohol-associated HCC tissues. TLR3 and ACTG1 were proposed as potential biomarkers of alcohol-associated HCC. Investigation into the regulatory functions of miR-6819-3p and miR-6877-3P may provide novel insights into the treatment of alcohol-associated HCC.

RUGOSE SPIRALING WHITEFLY-A NEW PEST OF OIL PALM IN INDIA

Rugose spiraling whitefly, Aleurodicus rugioperculatus Martin is found infesting oil palm plantation in the West Godavari district of Andhra Pradesh, India in 2017. Adults are tiny sized white flies with brown colour bands on the wings. Presence of wary filaments covering the eggs and immature stages on the undersurface of the leaves is the characteristic feature of identity. Secretion of honeydew and formation of black sooty mold layer on the lower leaves of same plant and or on other plants including weeds in the ground are good indication of the pest incidence. Coconut is the most preferred host plant followed by oil palm and other horticultural plants viz. banana, guava, heliconia, papaya while pineapple and palmyra are free from the pest incidence. Application of 0.1 per cent neem oil (I 0000 ppm) mixed with I per cent RIN detergent powder proved to be effective in suppressing the pest. Keywords: Rugose spiraling whitefly, oil palm, Andhra Pradesh, parasitoid, sooty mold.

OFD1 and VFL3/CCDC61 in basal body positioning and docking in Paramecium

Objectives Ciliogenesis is conditioned by a correct positioning/ anchoring of the basal body at the cell surface. In Paramecium we have shown that three conserved proteins FOR20, centrin 2 (CEN2) and centrin 3 (CEN3) participates in this process, with FOR20 and CEN2 being also involved in the transition zone assembly. We established a chronology in basal body assembly: CEN2 is required for FOR20 recruitment, the latter being necessary to recruit CEN3. Our goal now is to integrate others molecules in this cascade.

Centrin 2 is required for mouse olfactory ciliary trafficking and development of ependymal cilia planar polarity.

Centrins are ancient calmodulin-related Ca(2+)-binding proteins associated with basal bodies. In lower eukaryotes, Centrin2 (CETN2) is required for basal body replication and positioning, although its function in mammals is undefined. We generated a germline CETN2 knock-out (KO) mouse presenting with syndromic ciliopathy including dysosmia and hydrocephalus. Absence of CETN2 leads to olfactory cilia loss, impaired ciliary trafficking of olfactory signaling proteins, adenylate cyclase III (ACIII), and cyclic nucleotide-gated (CNG) channel, as well as disrupted basal body apical migration in postnatal olfactory sensory neurons (OSNs). In mutant OSNs, cilia base-anchoring of intraflagellar transport components IFT88, the kinesin-II subunit KIF3A, and cytoplasmic dynein 2 appeared compromised. Although the densities of mutant ependymal and respiratory cilia were largely normal, the planar polarity of mutant ependymal cilia was disrupted, resulting in uncoordinated flow of CSF. Transgenic expression of GFP-CETN2 rescued the Cetn2-deficiency phenotype. These results indicate that mammalian basal body replication and ciliogenesis occur independently of CETN2; however, mouse CETN2 regulates protein trafficking of olfactory cilia and participates in specifying planar polarity of ependymal cilia.

CETN1 is a cancer testis antigen with expression in prostate and pancreatic cancers

BackgroundThe Cancer Testis Antigens (CTAs) are a group of genes that are highly expressed in the normal testis and several types of cancer. Due to their restricted expression in normal adult tissues, CTAs have been attractive targets for immunotherapy and biomarker development. In this work, we discovered that Centrin 1 (CETN1) which is found in the centrosome of all eukaryotes, may be a member of this group and is highly expressed in prostate and pancreatic cancer. Three members of the centrin family of calcium binding proteins (CETN) are localized to the centrosome in all eukaryotes with CDC31 being the sole yeast homolog. CETN1 is a retrogene that probably arose from a retrotransposition of CETN2, an X-linked gene. A previous mouse study shows that CETN1 is expressed solely in the testis, while CETN2 is expressed in all organs.ResultsIn this work, we show that CETN1 is a new member of the growing group of CTAs. Through the mining of publicly available microarray data, we discovered that human CETN1 expression but not CETN2 or CETN3 is restricted to the testis. In fact, CETN1 is actually down-regulated in testicular malignancies compared to normal testis. Using q-PCR, CETN1 expression is shown to be highly up-regulated in cancer of the prostate and in pancreatic xenografts. Unexpectedly however, CETN1 expression was virtually absent in various cell lines until they were treated with the DNA demethylation agent 5’AZA-2’Deoxycytidine (AZA) but showed no increased expression upon incubation with Histone deacetylase inhibitor Trichostatin-A (TSA) alone. Additionally, like most CTAs, CETN1 appears to be an intrinsically disordered protein which implies that it may occupy a hub position in key protein interaction networks in cancer. Neither CETN1 nor CETN2 could compensate for loss of CDC31 expression in yeast which is analogous to published data for CETN3.ConclusionsThis work suggests that CETN1 is a novel CTA with expression in cancer of the prostate and pancreas. In cell lines, the expression is probably regulated by promoter methylation, while the method of regulation in normal adult tissues remains unknown.

Germline deletion of Cetn1 causes infertility in male mice

Centrins are calmodulin-like Ca(2+)-binding proteins that can be found in all ciliated eukaryotic cells from yeast to mammals. Expressed in male germ cells and photoreceptors, centrin 1 (CETN1) resides in the photoreceptor transition zone and connecting cilium. To identify its function in mammals, we deleted Cetn1 by homologous recombination. Cetn1(-/-) mice were viable and showed no sign of retina degeneration suggesting that CETN1 is nonessential for photoreceptor ciliogenesis or structural maintenance. Phototransduction components localized normally to the Cetn1(-/-) photoreceptor outer segments, and loss of CETN1 had no effect on light-induced translocation of transducin to the inner segment. Although Cetn1(-/-) females and Cetn1(+/-) males had normal fertility, Cetn1(-/-) males were infertile. The Cetn1(-/-) testes size was normal, and spermatogonia as well as spermatocytes developed normally. However, spermatids lacked tails suggesting severe defects at the late maturation phase of spermiogenesis. Viable sperm cells were absent and the few surviving spermatozoa were malformed. Light and electron microscopy analyses of Cetn1(-/-) spermatids revealed failures in centriole rearrangement during basal body maturation and in the basal-body-nucleus connection. These results confirm an essential role for CETN1 in late steps of spermiogenesis and spermatid maturation.

Comparative Genomic and Phylogenetic Analyses of Calcium ATPases and Calcium-Regulated Proteins in the Apicomplexa

The phylum Apicomplexa comprises a large group of early branching eukaryotes that includes a number of human and animal parasites. Calcium controls a number of vital processes in apicomplexans including protein secretion, motility, and differentiation. Despite the importance of calcium as a second messenger, very little is known about the systems that control homeostasis or that regulate calcium signaling in parasites. The recent completion of many apicomplexan genomes provides new opportunity to define calcium response pathways in this group of parasites in comparison to model organisms. Whole-genome comparison between the apicomplexans Plasmodium spp., Cryptosporidium spp., and Toxoplasma gondii revealed the presence of several P-Type Ca2+ transporting ATPases including a single endoplasmic reticulum (ER)-type sarcoplasmic-endoplasmic reticulum Ca2+ ATPase, several Golgi-like Ca2+ ATPases, and a single Ca2+/H+ exchanger. Only T. gondii showed evidence of plasma membrane-type Ca2+ ATPases or voltage-gated calcium channels. Despite pharmacological evidence for IP3 and ryanodine-mediated calcium release, animal-type calcium channels were not readily identified in parasites, indicating they are more similar to plants. Downstream of calcium release, a variety of EF-hand-containing proteins regulate calcium responses. Our analyses detected a single conserved calmodulin (CaM) homologue, 3 distinct centrin (CETN)-caltractin-like proteins, one of which is shared with ciliates, and a variety of deep-branching, CaM-CETN-like proteins. Apicomplexans were also found to contain a wide array of calcium-dependent protein kinases (CDPKs), which are commonly found in plants. Toxoplasma gondii contains more than 20 CDPK or CDPK-related kinases, which likely regulate a variety of responses including secretion, motility, and differentiation. Genomic and phylogenetic comparisons revealed that apicomplexans contain a variety of unusual calcium response pathways that are distinct from those seen in vertebrates. Notably, plant-like pathways for calcium release channels and calcium-dependent kinases are found in apicomplexans. The experimental flexibility of T. gondii should allow direct experimental manipulation of these pathways to validate their biological roles. The central importance of calcium in signaling and development, and the novel characteristics of many of these systems, indicates that parasite calcium pathways may be exploited as new therapeutic targets for intervention.

1H resonance assignment of the C-terminal domain of Human Centrin2 (LC-HsCen2: M84-Y172)

1H resonance assignment of the C-terminal domain of Human Centrin2 (LC-HsCen2: M84-Y172)

Centrosome Protein Centrin 2/Caltractin 1 Is Part of the Xeroderma Pigmentosum Group C Complex That Initiates Global Genome Nucleotide Excision Repair

Nucleotide excision repair (NER) is carried out by xeroderma pigmentosum (XP) factors. Before the excision reaction, DNA damage is recognized by a complex originally thought to contain the XP group C responsible gene product (XPC) and the human homologue of Rad23 B (HR23B). Here, we show that centrin 2/caltractin 1 (CEN2) is also a component of the XPC repair complex. We demonstrate that nearly all XPC complexes contain CEN2, that CEN2 interacts directly with XPC, and that CEN2, in cooperation with HR23B, stabilizes XPC, which stimulates XPC NER activity in vitro. CEN2 has been shown to play an important role in centrosome duplication. Thus, those findings suggest that the XPC-CEN2 interaction may reflect coupling of cell division and NER.