Biomedical subjects

Alexander Schleiffer

Publications and source records attributed to Alexander Schleiffer.

13 recordsLinked to original sources

PubMed2026

Systematic discovery of pathogen effector functions across human pathogens and pathways.

Pathogens deploy effector proteins to exploit host cell biology, and most effector open reading frames (ORFs) are rapidly evolving and lack functional annotation. We developed the effector ORFeome (eORFeome), a scalable functional genomics platform encompassing 3,835 effector ORFs from diverse viruses, bacteria, and parasites. High-throughput barcoded screens across nuclear factor κB (NF-κB), apoptosis, p53, cGAS-STING, and major histocompatibility complex class I (MHC class I) pathways revealed novel pathway-modulating functions for hundreds of uncharacterized eORFs, unexpected activities of known effectors, and distinct pathway-specific functions encoded by single ORFs. Illustrating the power of this approach, we identified HHV6A U14 as a p53 antagonist, HHV7 U21 as a dual-function STING antagonist and MHC-I antigen display inhibitor, and adenoviral 13.6K/i-leader protein as a de novo-evolved TAP inhibitor that suppresses MHC-I display. These results establish a general framework for systematic effector annotation, uncover new mechanisms of host-pathogen interaction across kingdoms, and highlight pathogen effectors as a versatile toolkit for rewiring and probing human cellular pathways.

Humans↗

PubMed2006

Wapl controls the dynamic association of cohesin with chromatin.

Cohesin establishes sister-chromatid cohesion from S phase until mitosis or meiosis. To allow chromosome segregation, cohesion has to be dissolved. In vertebrate cells, this process is mediated in part by the protease separase, which destroys a small amount of cohesin, but most cohesin is removed from chromosomes without proteolysis. How this is achieved is poorly understood. Here, we show that the interaction between cohesin and chromatin is controlled by Wapl, a protein implicated in heterochromatin formation and tumorigenesis. Wapl is associated with cohesin throughout the cell cycle, and its depletion blocks cohesin dissociation from chromosomes during the early stages of mitosis and prevents the resolution of sister chromatids until anaphase, which occurs after a delay. Wapl depletion also increases the residence time of cohesin on chromatin in interphase. Our data indicate that Wapl is required to unlock cohesin from a particular state in which it is stably bound to chromatin.

Amino Acid Sequence↗

PubMed2006

DOUTfinder--identification of distant domain outliers using subsignificant sequence similarity.

DOUTfinder is a web-based tool facilitating protein domain detection among related protein sequences in the twilight zone of sequence similarity. The sequence set required for this analysis can be provided by the user or will be collected using PSI-BLAST if a single sequence is given as an input. The obtained sequence family is analyzed for known Pfam and SMART domains, and the thereby identified subsignificant domain similarities are evaluated further. Domains with several subthreshold hits in the query set are ranked based on a sum-score function and likely homologous domains are suggested according to established cut-offs. By providing a post-filtering procedure for subsignificant domain hits DOUTfinder allows the detection of non-trivial domain relationships and can thereby lead to new insights into the function and evolution of distantly related sequence families. DOUTfinder is available at http://mendel.imp.ac.at/dout/.

Humans↗

PubMed2006

Human Scc4 is required for cohesin binding to chromatin, sister-chromatid cohesion, and mitotic progression.

BACKGROUND: Sister-chromatid cohesion depends on the cohesin complex whose association with chromatin is mediated by Scc2 and Scc4 in budding yeast. Both cohesin and Scc2 have been conserved from yeast to humans, but no Scc4 orthologs have been identified. Mutation of Scc2 orthologs causes defects in cohesion, transcription, and development, resulting in Cornelia de Lange syndrome in humans. RESULTS: We have identified a family of tetratricopeptide repeat proteins that share weak sequence similarities with yeast Scc4. This family includes MAU-2, which is required for development of the nervous system in Caenorhabditis elegans. We show that the human member of this family is associated with Scc2, is bound to chromatin from telophase until prophase, and is required for association of cohesin with chromatin during interphase. Cells lacking Scc4 lose sister-chromatid cohesion precociously and arrest in prometaphase. Mitotic chromosomes in Scc4-depleted cells lack cohesin, even though the cohesin-protecting proteins Sgo1 and Bub1 are normally enriched at centromeres and separase does not seem to be active. CONCLUSION: Our data indicate that human Scc4 is required for the association of cohesin with chromatin, which is a prerequisite for the establishment of sister-chromatid cohesion and for chromosome biorientation in mitosis. The proteinaceous machinery that is required for loading of cohesin onto chromatin is therefore conserved from yeast to humans. The finding that Caenorhabditis elegans MAU-2 is an ortholog of Scc4 further supports the notion that the Scc2-Scc4 complex is required for developmental processes in metazoans.

Amino Acid Sequence↗

PubMed2006

Mitotic activation of the kinase Aurora-A requires its binding partner Bora.

The protein kinase Aurora-A is required for centrosome maturation, spindle assembly, and asymmetric protein localization during mitosis. Here, we describe the identification of Bora, a conserved protein that is required for the activation of Aurora-A at the onset of mitosis. In the Drosophila peripheral nervous system, bora mutants have defects during asymmetric cell division identical to those observed in aurora-A. Furthermore, overexpression of bora can rescue defects caused by mutations in aurora-A. Bora is conserved in vertebrates, and both Drosophila and human Bora can bind to Aurora-A and activate the kinase in vitro. In interphase cells, Bora is a nuclear protein, but upon entry into mitosis, Bora is excluded from the nucleus and translocates into the cytoplasm in a Cdc2-dependent manner. We propose a model in which activation of Cdc2 initiates the release of Bora into the cytoplasm where it can bind and activate Aurora-A.

Animals↗

PubMed2005

Molecular processes during fat cell development revealed by gene expression profiling and functional annotation.

BACKGROUND: Large-scale transcription profiling of cell models and model organisms can identify novel molecular components involved in fat cell development. Detailed characterization of the sequences of identified gene products has not been done and global mechanisms have not been investigated. We evaluated the extent to which molecular processes can be revealed by expression profiling and functional annotation of genes that are differentially expressed during fat cell development. RESULTS: Mouse microarrays with more than 27,000 elements were developed, and transcriptional profiles of 3T3-L1 cells (pre-adipocyte cells) were monitored during differentiation. In total, 780 differentially expressed expressed sequence tags (ESTs) were subjected to in-depth bioinformatics analyses. The analysis of 3'-untranslated region sequences from 395 ESTs showed that 71% of the differentially expressed genes could be regulated by microRNAs. A molecular atlas of fat cell development was then constructed by de novo functional annotation on a sequence segment/domain-wise basis of 659 protein sequences, and subsequent mapping onto known pathways, possible cellular roles, and subcellular localizations. Key enzymes in 27 out of 36 investigated metabolic pathways were regulated at the transcriptional level, typically at the rate-limiting steps in these pathways. Also, coexpressed genes rarely shared consensus transcription-factor binding sites, and were typically not clustered in adjacent chromosomal regions, but were instead widely dispersed throughout the genome. CONCLUSIONS: Large-scale transcription profiling in conjunction with sophisticated bioinformatics analyses can provide not only a list of novel players in a particular setting but also a global view on biological processes and molecular networks.

3T3-L1 Cells↗

PubMed2005

Mammalian inscuteable regulates spindle orientation and cell fate in the developing retina.

During mammalian neurogenesis, progenitor cells can divide with the mitotic spindle oriented parallel or perpendicular to the surface of the neuroepithelium. Perpendicular divisions are more likely to be asymmetric and generate one progenitor and one neuronal precursor. Whether the orientation of the mitotic spindle actually determines their asymmetric outcome is unclear. Here, we characterize a mammalian homolog of Inscuteable (mInsc), a key regulator of spindle orientation in Drosophila. mInsc is expressed temporally and spatially in a manner that suggests a role in orienting the mitotic spindle in the developing nervous system. Using retroviral RNAi in rat retinal explants, we show that downregulation of mInsc inhibits vertical divisions. This results in enhanced proliferation, consistent with a higher frequency of symmetric divisions generating two proliferating cells. Our results suggest that the orientation of neural progenitor divisions is important for cell fate specification in the retina and determines their symmetric or asymmetric outcome.

Animals↗

PubMed2005

A REC8-dependent plant Shugoshin is required for maintenance of centromeric cohesion during meiosis and has no mitotic functions.

During meiosis, sequential release of sister chromatid cohesion (SSC) during two successive nuclear divisions allows the production of haploid gametes from diploid progenitor cells. Release of SSC along chromosome arms allows first a reductional segregation of homologs, and, subsequently, release of centromeric cohesion at anaphase II allows the segregation of chromatids. The Shugoshin (SGO) protein family plays a major role in the protection of centromeric cohesion in Drosophila and yeast. We have isolated a maize mutant that displays premature loss of centromeric cohesion at anaphase I. We showed that this phenotype is due to the absence of ZmSGO1 protein, a maize shugoshin homolog. We also show that ZmSGO1 is localized to the centromeres. The ZmSGO1 protein is not found on mitotic chromosomes and has no obvious mitotic function. On the basis of these results, we propose that ZmSGO1 specifically maintains centromeric cohesion during meiosis I and therefore suggest that SGO1 core functions during meiosis are conserved across kingdoms and in large-genome species. However, in contrast to other Shugoshins, we observed an early and REC8-dependent recruitment of ZmSGO1 in maize, suggesting that control of SGO1 recruitment to chromosomes is different in plants than in other model organisms.

Cell Cycle Proteins↗

PubMed2004

The Drosophila meiotic kleisin C(2)M functions before the meiotic divisions.

Stepwise and regionally controlled resolution of sister chromatid cohesion is thought to be crucial for faithful chromosome segregation during meiotic divisions. In yeast, the meiosis-specific alpha-kleisin subunit of the cohesin complex, Rec8, is protected from cleavage by separase but only during meiosis I and specifically within the pericentromeric region. While the Drosophila genome does not contain an obvious Rec8 orthologue, as other animal and plant genomes, it includes c(2)M, which encodes a distant alpha-kleisin family member involved in female meiosis. C(2)M associates in vivo with the Smc3 cohesin subunit, as previously shown for yeast Rec8. In contrast to Rec8, however, C(2)M accumulates predominantly after the pre-meiotic S-phase. Moreover, after association with the synaptonemal complex, it disappears again and cannot be detected on meiotic chromosomes by metaphase I. C(2)M cleavage fragments are not observed during completion of the meiotic divisions, and mutations within putative separase cleavage sites do not interfere with meiotic chromosome segregation. Therefore, C(2)M appears to function within the synaptonemal complex during prophase I but possibly not thereafter. This suggests that C(2)M may not confer sister chromatid cohesion needed for meiosis I and II chromosome segregation.

Animals↗

PubMed2004

MYRbase: analysis of genome-wide glycine myristoylation enlarges the functional spectrum of eukaryotic myristoylated proteins.

We evaluated the evolutionary conservation of glycine myristoylation within eukaryotic sequences. Our large-scale cross-genome analyses, available as MYRbase, show that the functional spectrum of myristoylated proteins is currently largely underestimated. We give experimental evidence for in vitro myristoylation of selected predictions. Furthermore, we classify five membrane-attachment factors that occur most frequently in combination with, or even replacing, myristoyl anchors, as some protein family examples show.

Amino Acid Sequence↗

PubMed2004

From a single double helix to paired double helices and back.

The propagation of our genomes during cell proliferation depends on the movement of sister DNA molecules produced by DNA replication to opposite sides of the cell before it divides. This feat is achieved by microtubules in eukaryotic cells but it has long remained a mystery how cells ensure that sister DNAs attach to microtubules with opposite orientations, known as amphitelic attachment. It is currently thought that sister chromatid cohesion has a crucial role. By resisting the forces exerted by microtubules, sister chromatid cohesion gives rise to tension that is thought essential for stabilizing kinetochore-microtubule attachments. Efficient amphitelic attachment is therefore achieved by an error correction mechanism that selectively eliminates connections that do not give rise to tension. Cohesion between sister chromatids is mediated by a multisubunit complex called cohesin which forms a gigantic ring structure. It has been proposed that sister DNAs are held together owing to their becoming entrapped within a single cohesin ring. Cohesion between sister chromatids is destroyed at the metaphase to anaphase transition by proteolytic cleavage of cohesin's Scc1 subunit by a thiol protease called separase, which severs the ring and thereby releases sister DNAs.

Cell Cycle Proteins↗

PubMed2003

Kleisins: a superfamily of bacterial and eukaryotic SMC protein partners.

We describe a superfamily of eukaryotic and prokaryotic proteins (kleisins) that includes ScpA, Scc1, Rec8, and Barren. Scc1 interacts with SMC proteins through N- and C-terminal domains to form a ring-like structure. Since these are the only domains conserved among kleisins, we suggest that ring formation with SMC proteins may define this family.

Adenosine Triphosphatases↗

PubMed2002

Eco1 is a novel acetyltransferase that can acetylate proteins involved in cohesion.

Cohesion between sister chromatids is established during S phase and maintained through G2 phase until it is resolved in anaphase (for review, see [1-3]). In Saccharomyces cerevisiae, a complex consisting of Scc1, Smc1, Smc3, and Scc3 proteins, called "cohesin," mediates the connection between sister chromatids. The evolutionary conserved yeast protein Eco1 is required for establishment of sister chromatid cohesion during S phase but not for its further maintenance during G2 or M phases or for loading the cohesin complex onto DNA. We address the molecular functions of Eco1 with sensitive sequence analytic techniques, including hidden Markov model domain fragment searches. We found a two-domain architecture with an N-terminal C2H2 Zn finger-like domain and an approximately 150 residue C-terminal domain with an apparent acetyl coenzyme A binding motif (http://mendel.imp.univie.ac.at/SEQUENCES/ECO1/). Biochemical tests confirm that Eco1 has the acetyltransferase activity in vitro. In vitro Eco1 acetylates itself and components of the cohesin complex but not histones. Thus, the establishment of cohesion between sister chromatids appears to be regulated, directly or indirectly, by a specific acetyltransferase.

Acetylation↗