Interplay of Substrate Retention and Export Signals in Endoplasmic Reticulum Quality Control

BACKGROUND: Endoplasmic reticulum (ER) quality control mechanisms are part of a comprehensive system to manage cell stress. The flux of molecules is monitored to retain folding intermediates and target misfolded molecules to ER-associated degradation (ERAD) pathways. The mechanisms of sorting remain unclear. While some proteins are retained statically, the classical model substrate CPY* is found in COPII transport vesicles, suggesting a retrieval mechanism for retention. However, its management can be even more dynamic. If ERAD is saturated under stress, excess CPY* traffics to the vacuole for degradation. These observations suggest that misfolded proteins might display different signals for their management. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report the existence of a functional ER exit signal in the pro-domain of CPY*. Compromising its integrity causes ER retention through exclusion from COPII vesicles. The signal co-exists with other signals used for retention and degradation. Physiologically, the export signal is important for stress tolerance. Disabling it converts a benign protein into one that is intrinsically cytotoxic. CONCLUSIONS/SIGNIFICANCE: These data reveal the remarkable interplay between opposing signals embedded within ERAD substrate molecules and the mechanisms that decipher them. Our findings demonstrate the diversity of mechanisms deployed for protein quality control and maintenance of protein homeostasis

A DNA Polymerase α Accessory Protein, Mcl1, Is Required for Propagation of Centromere Structures in Fission Yeast

Specialized chromatin exists at centromeres and must be precisely transmitted during DNA replication. The mechanisms involved in the propagation of these structures remain elusive. Fission yeast centromeres are composed of two chromatin domains: the central CENP-ACnp1 kinetochore domain and flanking heterochromatin domains. Here we show that fission yeast Mcl1, a DNA polymerase α (Polα) accessory protein, is critical for maintenance of centromeric chromatin. In a screen for mutants that alleviate both central domain and outer repeat silencing, we isolated several cos mutants, of which cos1 is allelic to mcl1. The mcl1-101 mutation causes reduced CENP-ACnp1 in the central domain and an aberrant increase in histone acetylation in both domains. These phenotypes are also observed in a mutant of swi7+, which encodes a catalytic subunit of Polα. Mcl1 forms S-phase-specific nuclear foci, which colocalize with those of PCNA and Polα. These results suggest that Mcl1 and Polα are required for propagation of centromere chromatin structures during DNA replication

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency–Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research

Distinct roles for antiparallel microtubule pairing and overlap during early spindle assembly

During spindle assembly, microtubules may attach to kinetochores or pair to form antiparallel pairs or interpolar microtubules, which span the two spindle poles and contribute to mitotic pole separation and chromosome segregation. Events in the specification of the interpolar microtubules are poorly understood. Using three-dimensional electron tomography and analysis of spindle dynamical behavior in living cells, we investigated the process of spindle assembly. Unexpectedly, we found that the phosphorylation state of an evolutionarily conserved Cdk1 site (S360) in γ-tubulin is correlated with the number and organization of interpolar microtubules. Mimicking S360 phosphorylation (S360D) results in bipolar spindles with a normal number of microtubules but lacking interpolar microtubules. Inhibiting S360 phosphorylation (S360A) results in spindles with interpolar microtubules and high-angle, antiparallel microtubule pairs. The latter are also detected in wild-type spindles <1 μm in length, suggesting that high-angle microtubule pairing represents an intermediate step in interpolar microtubule formation. Correlation of spindle architecture with dynamical behavior suggests that microtubule pairing is sufficient to separate the spindle poles, whereas interpolar microtubules maintain the velocity of pole displacement during early spindle assembly. Our findings suggest that the number of interpolar microtubules formed during spindle assembly is controlled in part through activities at the spindle poles