Distinct patterns of proteostasis network gene expression are associated with different prognoses in melanoma patients.

The proteostasis network (PN) is a collection of protein folding and degradation pathways that spans cellular compartments and acts to preserve the integrity of the proteome. The differential expression of PN genes is a hallmark of many cancers, and the inhibition of protein quality control factors is an effective way to slow cancer cell growth. However, little is known about how the expression of PN genes differs between patients and how this impacts survival outcomes. To address this, we applied unbiased hierarchical clustering to gene expression data obtained from primary and metastatic cutaneous melanoma (CM) samples and found that two distinct groups of individuals emerge across each sample type. These patient groups are distinguished by the differential expression of genes encoding ATP-dependent and ATP-independent chaperones, and proteasomal subunits. Differences in PN gene expression were associated with increased levels of the transcription factors, MEF2A, SP4, ZFX, CREB1 and ATF2, as well as markedly different survival outcomes. However, surprisingly, similar PN alterations in primary and metastatic samples were associated with discordant survival outcomes in patients. Our findings reveal that the expression of PN genes demarcates CM patients and highlights several new proteostasis sub-networks that could be targeted for more effective suppression of CM within specific individuals.

Loss of MTCH-1 suppresses age-related proteostasis collapse through the inhibition of programmed cell death factors.

The age-related loss of protein homeostasis (proteostasis) is at the heart of numerous neurodegenerative diseases. Therefore, finding ways to preserve proteome integrity in aged cells may be a powerful way to promote long-term health. Here, we show that reducing the activity of a highly conserved mitochondrial outer membrane protein, MTCH-1/MTCH2, suppresses age-related proteostasis collapse in Caenorhabditis elegans without disrupting development, growth, or reproduction. Loss of MTCH-1 does not influence proteostasis capacity in aged tissues through previously described pathways but instead operates by reducing CED-4 levels. This results in the sequestration of HSP-90 by inactive CED-3, which in turn leads to an increase in HSF-1 activity, transcriptional remodeling of the proteostasis network, and maintenance of proteostasis capacity with age. Together, our findings reveal a role for programmed cell death factors in determining proteome health and suggest that inhibiting MTCH-1 activity in adulthood may safeguard the aging proteome and suppress age-related diseases.

Nascent RNA antagonizes the interaction of a set of regulatory proteins with chromatin.

A number of regulatory factors are recruited to chromatin by specialized RNAs. Whether RNA has a more general role in regulating the interaction of proteins with chromatin has not been determined. We used proteomics methods to measure the global impact of nascent RNA on chromatin in embryonic stem cells. Surprisingly, we found that nascent RNA primarily antagonized the interaction of chromatin modifiers and transcriptional regulators with chromatin. Transcriptional inhibition and RNA degradation induced recruitment of a set of transcriptional regulators, chromatin modifiers, nucleosome remodelers, and regulators of higher-order structure. RNA directly bound to factors, including BAF, NuRD, EHMT1, and INO80 and inhibited their interaction with nucleosomes. The transcriptional elongation factor P-TEFb directly bound pre-mRNA, and its recruitment to chromatin upon Pol II inhibition was regulated by the 7SK ribonucleoprotein complex. We postulate that by antagonizing the interaction of regulatory proteins with chromatin, nascent RNA links transcriptional output with chromatin composition.

An optimised saliva collection method to produce high-yield, high-quality RNA for translational research.

Saliva represents an ideal matrix for diagnostic biomarker development as it is readily available and requires no invasive collection procedures. However, salivary RNA is labile and rapidly degrades. Previous attempts to isolate RNA from saliva have yielded poor quality and low concentrations. Here we compare collection and processing methods and propose an approach for future studies. The effects of RNA stabilisers, storage temperatures, length of storage and fasting windows were investigated on pooled saliva samples from healthy volunteers. Isolated RNA was assessed for concentration and quality. Bacterial growth was investigated through RT-PCR using bacterial and human primers. Optimal conditions were implemented and quality controlled in a clinical setting. The addition of RNAlater increased mean RNA yield from 4912 ng/µl to 15,473 ng and RNA Integrity Number (RIN) from 4.5 to 7.0. No significant changes to RNA yield were observed for storage at room temperature beyond 1 day or at -80 °C. Bacterial growth did not occur in samples stored at ambient temperature for up to a week. There was a trend towards higher RNA concentration when saliva was collected after overnight fasting but no effect on RIN. In the clinic, RNA yields of 6307 ng and RINs of 3.9 were achieved, improving on previous reports. The method we describe here is a robust, clinically feasible saliva collection method using preservative that gives high concentrations and improved RINs compared to saliva collected without preservative.

Narrative ability of children with speech sound disorders and the prediction of later literacy skills.

PURPOSE: The main purpose of this study was to examine how children with isolated speech sound disorders (SSDs; n = 20), children with combined SSDs and language impairment (LI; n = 20), and typically developing children (n = 20), ages 3;3 (years;months) to 6;6, differ in narrative ability. The second purpose was to determine if early narrative ability predicts school-age (8-12 years) literacy skills. METHOD: This study employed a longitudinal cohort design. The children completed a narrative retelling task before their formal literacy instruction began. The narratives were analyzed and compared for group differences. Performance on these early narratives was then used to predict the children's reading decoding, reading comprehension, and written language ability at school age. RESULTS: Significant group differences were found in children's (a) ability to answer questions about the story, (b) use of story grammars, and (c) number of correct and irrelevant utterances. Regression analysis demonstrated that measures of story structure and accuracy were the best predictors of the decoding of real words, reading comprehension, and written language. Measures of syntax and lexical diversity were the best predictors of the decoding of nonsense words. CONCLUSION: Combined SSDs and LI, and not isolated SSDs, impact a child's narrative abilities. Narrative retelling is a useful task for predicting which children may be at risk for later literacy problems.