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Roles of the developmental regulator unc-62/Homothorax in limiting longevity in Caenorhabditis elegans.

Van Nostrand EL, Sánchez-Blanco A, Wu B, Nguyen A, Kim SK - PLoS Genet. (2013)

Bottom Line: Through analysis of the downstream consequences of unc-62 knockdown, we identify multiple effects linked to aging.Second, unc-62 RNAi results in a broad increase in expression of intestinal genes that typically decrease expression with age, suggesting that unc-62 activity balances intestinal resource allocation between yolk protein expression and fertility on the one hand and somatic functions on the other.These results illustrate how unc-62 regulation of intestinal gene expression is responsible for limiting lifespan during the normal aging process.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Stanford University Medical Center, Stanford, California, USA.

ABSTRACT
The normal aging process is associated with stereotyped changes in gene expression, but the regulators responsible for these age-dependent changes are poorly understood. Using a novel genomics approach, we identified HOX co-factor unc-62 (Homothorax) as a developmental regulator that binds proximal to age-regulated genes and modulates lifespan. Although unc-62 is expressed in diverse tissues, its functions in the intestine play a particularly important role in modulating lifespan, as intestine-specific knockdown of unc-62 by RNAi increases lifespan. An alternatively-spliced, tissue-specific isoform of unc-62 is expressed exclusively in the intestine and declines with age. Through analysis of the downstream consequences of unc-62 knockdown, we identify multiple effects linked to aging. First, unc-62 RNAi decreases the expression of yolk proteins (vitellogenins) that aggregate in the body cavity in old age. Second, unc-62 RNAi results in a broad increase in expression of intestinal genes that typically decrease expression with age, suggesting that unc-62 activity balances intestinal resource allocation between yolk protein expression and fertility on the one hand and somatic functions on the other. Finally, in old age, the intestine shows increased expression of several aberrant genes; these UNC-62 targets are expressed predominantly in neuronal cells in developing animals, but surprisingly show increased expression in the intestine of old animals. Intestinal expression of some of these genes during aging is detrimental for longevity; notably, increased expression of insulin ins-7 limits lifespan by repressing activity of insulin pathway response factor DAF-16/FOXO in aged animals. These results illustrate how unc-62 regulation of intestinal gene expression is responsible for limiting lifespan during the normal aging process.

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UNC-62 RNAi decreases expression of collagen genes and increases expression of intestinal genes.(A) unc-62 RNAi decreases expression of collagen genes. Rows correspond to 77 collagen genes profiled in both our RNA-seq experiment as well as microarray studies of aging. Each row shows data for an individual collagen gene. (left) Color indicates RNA-seq fold-change between unc-62 RNAi and control, averaged from triplicate biological experiments. (right) The four columns indicate expression at day 2, 5, 8, and 11 from whole-worm microarrays (normalized to day 2 expression) (data from [10]). (B) Of the 99 genes down-regulated upon unc-62 RNAi (at 10% FDR) profiled by aging microarrays, 38 also decrease with age (3.9-fold enriched, p<10−20 by Fisher's exact test). (C) The 48 genes that increase in expression upon unc-62 RNAi (10% FDR) show a significant overlap with the 556 genes that decrease in expression with age (p<10−4) [10]. (D) unc-62 RNAi causes a broad increase in expression of intestinal genes. The histogram indicates the distribution of genes by average fold-change upon unc-62 RNAi. For 1699 genes with intestine-enriched expression [23], [33], [34] as well as the subset of 291 genes that also decrease expression with age [10], we determined the average mean-centered fold-change from triplicate RNA-seq experiments of unc-62 RNAi as compared to controls. Both intestine-enriched genes (orange) and intestine-enriched genes that decrease expression with age (pink) are significantly shifted towards increased expression upon unc-62 RNAi (p-value<10−20 by Kolmogorov-Smirnov test). (insert) The percent of genes with fold-change of 1.25 or more are indicated. (E) (left) An OPT-2:GFP reporter was used to measure intestinal morphology. In young adults, opt-2 localizes to the apical membrane of the intestine (class A). This structure deteriorates in older worms, as some worms contain the structure through only a portion of their body (class B) and others lack opt-2 expression altogether (class C). (right) Worms were placed on either unc-62 or control RNAi at day 1 of adulthood, and ∼30 worms for each were imaged and annotated at day 3 and day 6. Stacked bars indicate the percent of worms that were observed as class A (light grey), class B (dark grey), or class C (black).
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pgen-1003325-g006: UNC-62 RNAi decreases expression of collagen genes and increases expression of intestinal genes.(A) unc-62 RNAi decreases expression of collagen genes. Rows correspond to 77 collagen genes profiled in both our RNA-seq experiment as well as microarray studies of aging. Each row shows data for an individual collagen gene. (left) Color indicates RNA-seq fold-change between unc-62 RNAi and control, averaged from triplicate biological experiments. (right) The four columns indicate expression at day 2, 5, 8, and 11 from whole-worm microarrays (normalized to day 2 expression) (data from [10]). (B) Of the 99 genes down-regulated upon unc-62 RNAi (at 10% FDR) profiled by aging microarrays, 38 also decrease with age (3.9-fold enriched, p<10−20 by Fisher's exact test). (C) The 48 genes that increase in expression upon unc-62 RNAi (10% FDR) show a significant overlap with the 556 genes that decrease in expression with age (p<10−4) [10]. (D) unc-62 RNAi causes a broad increase in expression of intestinal genes. The histogram indicates the distribution of genes by average fold-change upon unc-62 RNAi. For 1699 genes with intestine-enriched expression [23], [33], [34] as well as the subset of 291 genes that also decrease expression with age [10], we determined the average mean-centered fold-change from triplicate RNA-seq experiments of unc-62 RNAi as compared to controls. Both intestine-enriched genes (orange) and intestine-enriched genes that decrease expression with age (pink) are significantly shifted towards increased expression upon unc-62 RNAi (p-value<10−20 by Kolmogorov-Smirnov test). (insert) The percent of genes with fold-change of 1.25 or more are indicated. (E) (left) An OPT-2:GFP reporter was used to measure intestinal morphology. In young adults, opt-2 localizes to the apical membrane of the intestine (class A). This structure deteriorates in older worms, as some worms contain the structure through only a portion of their body (class B) and others lack opt-2 expression altogether (class C). (right) Worms were placed on either unc-62 or control RNAi at day 1 of adulthood, and ∼30 worms for each were imaged and annotated at day 3 and day 6. Stacked bars indicate the percent of worms that were observed as class A (light grey), class B (dark grey), or class C (black).

Mentions: To further understand the roles of unc-62 during aging, we analyzed the 182 transcripts that show altered expression in unc-62 RNAi-treated worms, consisting of 67 transcripts that increase expression and 115 transcripts that decrease expression. The 115 targets that are (directly or indirectly) activated by wild-type unc-62 show a strong enrichment for genes specifically expressed in the hypodermis (2.6-fold; p<10−5). Notably, 41 of these 115 genes are collagen genes, out of a total of 90 collagen genes quantified in the RNA-seq experiment (29-fold enriched, p<10−15)(Figure 6A). These unc-62-activated genes also include 38 that decrease in expression during normal aging (a 3.9-fold enrichment, p<10−20) [10], including 26 of the 41 unc-62-activated collagen genes (Figure 6A–6B). These findings suggest that changes in gene expression of hypodermal genes upon unc-62 knockdown (particularly, collagen genes) are similar to changes normally observed as adult worms age; i.e., unc-62 RNAi mimics the normal aging process in the hypodermis.


Roles of the developmental regulator unc-62/Homothorax in limiting longevity in Caenorhabditis elegans.

Van Nostrand EL, Sánchez-Blanco A, Wu B, Nguyen A, Kim SK - PLoS Genet. (2013)

UNC-62 RNAi decreases expression of collagen genes and increases expression of intestinal genes.(A) unc-62 RNAi decreases expression of collagen genes. Rows correspond to 77 collagen genes profiled in both our RNA-seq experiment as well as microarray studies of aging. Each row shows data for an individual collagen gene. (left) Color indicates RNA-seq fold-change between unc-62 RNAi and control, averaged from triplicate biological experiments. (right) The four columns indicate expression at day 2, 5, 8, and 11 from whole-worm microarrays (normalized to day 2 expression) (data from [10]). (B) Of the 99 genes down-regulated upon unc-62 RNAi (at 10% FDR) profiled by aging microarrays, 38 also decrease with age (3.9-fold enriched, p<10−20 by Fisher's exact test). (C) The 48 genes that increase in expression upon unc-62 RNAi (10% FDR) show a significant overlap with the 556 genes that decrease in expression with age (p<10−4) [10]. (D) unc-62 RNAi causes a broad increase in expression of intestinal genes. The histogram indicates the distribution of genes by average fold-change upon unc-62 RNAi. For 1699 genes with intestine-enriched expression [23], [33], [34] as well as the subset of 291 genes that also decrease expression with age [10], we determined the average mean-centered fold-change from triplicate RNA-seq experiments of unc-62 RNAi as compared to controls. Both intestine-enriched genes (orange) and intestine-enriched genes that decrease expression with age (pink) are significantly shifted towards increased expression upon unc-62 RNAi (p-value<10−20 by Kolmogorov-Smirnov test). (insert) The percent of genes with fold-change of 1.25 or more are indicated. (E) (left) An OPT-2:GFP reporter was used to measure intestinal morphology. In young adults, opt-2 localizes to the apical membrane of the intestine (class A). This structure deteriorates in older worms, as some worms contain the structure through only a portion of their body (class B) and others lack opt-2 expression altogether (class C). (right) Worms were placed on either unc-62 or control RNAi at day 1 of adulthood, and ∼30 worms for each were imaged and annotated at day 3 and day 6. Stacked bars indicate the percent of worms that were observed as class A (light grey), class B (dark grey), or class C (black).
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pgen-1003325-g006: UNC-62 RNAi decreases expression of collagen genes and increases expression of intestinal genes.(A) unc-62 RNAi decreases expression of collagen genes. Rows correspond to 77 collagen genes profiled in both our RNA-seq experiment as well as microarray studies of aging. Each row shows data for an individual collagen gene. (left) Color indicates RNA-seq fold-change between unc-62 RNAi and control, averaged from triplicate biological experiments. (right) The four columns indicate expression at day 2, 5, 8, and 11 from whole-worm microarrays (normalized to day 2 expression) (data from [10]). (B) Of the 99 genes down-regulated upon unc-62 RNAi (at 10% FDR) profiled by aging microarrays, 38 also decrease with age (3.9-fold enriched, p<10−20 by Fisher's exact test). (C) The 48 genes that increase in expression upon unc-62 RNAi (10% FDR) show a significant overlap with the 556 genes that decrease in expression with age (p<10−4) [10]. (D) unc-62 RNAi causes a broad increase in expression of intestinal genes. The histogram indicates the distribution of genes by average fold-change upon unc-62 RNAi. For 1699 genes with intestine-enriched expression [23], [33], [34] as well as the subset of 291 genes that also decrease expression with age [10], we determined the average mean-centered fold-change from triplicate RNA-seq experiments of unc-62 RNAi as compared to controls. Both intestine-enriched genes (orange) and intestine-enriched genes that decrease expression with age (pink) are significantly shifted towards increased expression upon unc-62 RNAi (p-value<10−20 by Kolmogorov-Smirnov test). (insert) The percent of genes with fold-change of 1.25 or more are indicated. (E) (left) An OPT-2:GFP reporter was used to measure intestinal morphology. In young adults, opt-2 localizes to the apical membrane of the intestine (class A). This structure deteriorates in older worms, as some worms contain the structure through only a portion of their body (class B) and others lack opt-2 expression altogether (class C). (right) Worms were placed on either unc-62 or control RNAi at day 1 of adulthood, and ∼30 worms for each were imaged and annotated at day 3 and day 6. Stacked bars indicate the percent of worms that were observed as class A (light grey), class B (dark grey), or class C (black).
Mentions: To further understand the roles of unc-62 during aging, we analyzed the 182 transcripts that show altered expression in unc-62 RNAi-treated worms, consisting of 67 transcripts that increase expression and 115 transcripts that decrease expression. The 115 targets that are (directly or indirectly) activated by wild-type unc-62 show a strong enrichment for genes specifically expressed in the hypodermis (2.6-fold; p<10−5). Notably, 41 of these 115 genes are collagen genes, out of a total of 90 collagen genes quantified in the RNA-seq experiment (29-fold enriched, p<10−15)(Figure 6A). These unc-62-activated genes also include 38 that decrease in expression during normal aging (a 3.9-fold enrichment, p<10−20) [10], including 26 of the 41 unc-62-activated collagen genes (Figure 6A–6B). These findings suggest that changes in gene expression of hypodermal genes upon unc-62 knockdown (particularly, collagen genes) are similar to changes normally observed as adult worms age; i.e., unc-62 RNAi mimics the normal aging process in the hypodermis.

Bottom Line: Through analysis of the downstream consequences of unc-62 knockdown, we identify multiple effects linked to aging.Second, unc-62 RNAi results in a broad increase in expression of intestinal genes that typically decrease expression with age, suggesting that unc-62 activity balances intestinal resource allocation between yolk protein expression and fertility on the one hand and somatic functions on the other.These results illustrate how unc-62 regulation of intestinal gene expression is responsible for limiting lifespan during the normal aging process.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Stanford University Medical Center, Stanford, California, USA.

ABSTRACT
The normal aging process is associated with stereotyped changes in gene expression, but the regulators responsible for these age-dependent changes are poorly understood. Using a novel genomics approach, we identified HOX co-factor unc-62 (Homothorax) as a developmental regulator that binds proximal to age-regulated genes and modulates lifespan. Although unc-62 is expressed in diverse tissues, its functions in the intestine play a particularly important role in modulating lifespan, as intestine-specific knockdown of unc-62 by RNAi increases lifespan. An alternatively-spliced, tissue-specific isoform of unc-62 is expressed exclusively in the intestine and declines with age. Through analysis of the downstream consequences of unc-62 knockdown, we identify multiple effects linked to aging. First, unc-62 RNAi decreases the expression of yolk proteins (vitellogenins) that aggregate in the body cavity in old age. Second, unc-62 RNAi results in a broad increase in expression of intestinal genes that typically decrease expression with age, suggesting that unc-62 activity balances intestinal resource allocation between yolk protein expression and fertility on the one hand and somatic functions on the other. Finally, in old age, the intestine shows increased expression of several aberrant genes; these UNC-62 targets are expressed predominantly in neuronal cells in developing animals, but surprisingly show increased expression in the intestine of old animals. Intestinal expression of some of these genes during aging is detrimental for longevity; notably, increased expression of insulin ins-7 limits lifespan by repressing activity of insulin pathway response factor DAF-16/FOXO in aged animals. These results illustrate how unc-62 regulation of intestinal gene expression is responsible for limiting lifespan during the normal aging process.

Show MeSH