Gene expression profiles associated with aging and mortality in humans

We investigated the hypothesis that gene expression profiles in cultured cell lines from adults, aged 57–97 years, contain information about the biological age and potential longevity of the donors. We studied 104 unrelated grandparents from 31 Utah CEU (Centre d’Etude du Polymorphisme Humain – Utah) families, for whom lymphoblastoid cell lines were established in the 1980s. Combining publicly available gene expression data from these cell lines, and survival data from the Utah Population Database, we tested the relationship between expression of 2151 always-expressed genes, age, and survival of the donors. Approximately 16% of 2151 expression levels were associated with donor age: 10% decreased in expression with age, and 6% increased with age. Cell division cycle 42 (CDC42) and CORO1A exhibited strong associations both with age at draw and survival after draw (multiple comparisons-adjusted Monte Carlo P-value < 0.05). In general, gene expressions that increased with age were associated with increased mortality. Gene expressions that decreased with age were generally associated with reduced mortality. A multivariate estimate of biological age modeled from expression data was dominated by CDC42 expression, and was a significant predictor of survival after blood draw. A multivariate model of survival as a function of gene expression was dominated by CORO1A expression. This model accounted for approximately 23% of the variation in survival among the CEU grandparents. Some expression levels were negligibly associated with age in this cross-sectional dataset, but strongly associated with inter-individual differences in survival. These observations may lead to new insights regarding the genetic contribution to exceptional longevity

Concordant Gene Expression in Leukemia Cells and Normal Leukocytes Is Associated with Germline cis-SNPs

The degree to which gene expression covaries between different primary tissues within an individual is not well defined. We hypothesized that expression that is concordant across tissues is more likely influenced by genetic variability than gene expression which is discordant between tissues. We quantified expression of 11,873 genes in paired samples of primary leukemia cells and normal leukocytes from 92 patients with acute lymphoblastic leukemia (ALL). Genetic variation at >500,000 single nucleotide polymorphisms (SNPs) was also assessed. The expression of only 176/11,783 (1.5%) genes was correlated (p<0.008, FDR = 25%) in the two tissue types, but expression of a high proportion (20 of these 176 genes) was significantly related to cis-SNP genotypes (adjusted p<0.05). In an independent set of 134 patients with ALL, 14 of these 20 genes were validated as having expression related to cis-SNPs, as were 9 of 20 genes in a second validation set of HapMap cell lines. Genes whose expression was concordant among tissue types were more likely to be associated with germline cis-SNPs than genes with discordant expression in these tissues; genes affected were involved in housekeeping functions (GSTM2, GAPDH and NCOR1) and purine metabolism

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Generation of Transgenic Self-Incompatible Arabidopsis thaliana Shows a Genus-Specific Preference for Self-Incompatibility Genes

Brassicaceae species employ both self-compatibility and self-incompatibility systems to regulate post-pollination events. Arabidopsis halleri is strictly self-incompatible, while the closely related Arabidopsis thaliana has transitioned to self-compatibility with the loss of functional S-locus genes during evolution. The downstream signaling protein, ARC1, is also required for the self-incompatibility response in some Arabidopsis and Brassica species, and its gene is deleted in the A. thaliana genome. In this study, we attempted to reconstitute the SCR-SRK-ARC1 signaling pathway to restore self-incompatibility in A. thaliana using genes from A. halleri and B. napus, respectively. Several of the transgenic A. thaliana lines expressing the A. halleri SCR13-SRK13-ARC1 transgenes displayed self-incompatibility, while all the transgenic A. thaliana lines expressing the B. napus SCR1-SRK1-ARC1 transgenes failed to show any self-pollen rejection. Furthermore, our results showed that the intensity of the self-incompatibility response in transgenic A. thaliana plants was not associated with the expression levels of the transgenes. Thus, this suggests that there are differences between the Arabidopsis and Brassica self-incompatibility signaling pathways, which perhaps points to the existence of other factors downstream of B. napus SRK that are absent in Arabidopsis species