Limits...
Use of Drosophila as an evaluation method reveals imp as a candidate gene for type 2 diabetes in rat locus Niddm22.

Kawasaki K, Yamada S, Ogata K, Saito Y, Takahama A, Yamada T, Matsumoto K, Kose H - J Diabetes Res (2015)

Bottom Line: In the mutant, the expression of two of the insulin-like peptides encoded in the fly genome, dilp2 and dilp3, were found to be downregulated.Consistent with previous reports of dilp mutants, the imp mutant flies exhibited an extension of life span; in contrast, starvation tolerance was reduced.These results further reinforce the possibility that imp is involved in sugar metabolism by modulating insulin expression.

View Article: PubMed Central - PubMed

Affiliation: Division of Natural Sciences, Department of Life Science, International Christian University, Mitaka, Tokyo 181-8585, Japan.

ABSTRACT
Type 2 diabetes (T2D) is one of the most common human diseases. QTL analysis of the diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats has identified numerous hyperglycemic loci. However, molecular characterization and/or gene identification largely remains to be elucidated due mostly to the weak genetic variances contributed by each locus. Here we utilized Drosophila melanogaster as a secondary model organism for functional evaluation of the candidate gene. We demonstrate that the tissue specific knockdown of a homologue of igf2bp2 RNA binding protein leads to increased sugar levels similar to that found in the OLETF rat. In the mutant, the expression of two of the insulin-like peptides encoded in the fly genome, dilp2 and dilp3, were found to be downregulated. Consistent with previous reports of dilp mutants, the imp mutant flies exhibited an extension of life span; in contrast, starvation tolerance was reduced. These results further reinforce the possibility that imp is involved in sugar metabolism by modulating insulin expression.

Show MeSH

Related in: MedlinePlus

Expression pattern of the GFP-tagged protein trap. imp expression in third instar larva was examined using a protein trap strain ZCL0310, which expresses GFP-Imp chimeric protein in endogenous imp-expressing tissues. Bright field (a, c) and intrinsic GFP (b, d) images are shown. The GFP signal was detected in the central nervous system (CNS) (a, b), but only in a subset of neurons within the CNS. In contrast, GFP-Imp was not confirmed in the fat body (c, d). Bar, 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4363715&req=5

fig1: Expression pattern of the GFP-tagged protein trap. imp expression in third instar larva was examined using a protein trap strain ZCL0310, which expresses GFP-Imp chimeric protein in endogenous imp-expressing tissues. Bright field (a, c) and intrinsic GFP (b, d) images are shown. The GFP signal was detected in the central nervous system (CNS) (a, b), but only in a subset of neurons within the CNS. In contrast, GFP-Imp was not confirmed in the fat body (c, d). Bar, 100 μm.

Mentions: Previous studies showed that imp is expressed in the central nervous system and pole cells during embryonic development and germ cells in adults [26–28]. In order to examine the expression pattern of imp in larvae, we analyzed a protein trap strain, ZCL0310 [28]. In the third instar wandering larvae, the expression was exclusively detected in the central nervous system (CNS) (Figure 1). In contrast, imp is not expressed in other metabolically crucial tissues, including body wall muscle, fat body, gut, and oenocytes (Figure 1). Next we produced an CNS specific imp knockdown strain. The mutant had normal hatching rate and developmental growth. No morphological defect was observed. We confirmed that, in the third instar larvae, imp expression was reduced to about 20% of that of control (Figure 2(a)). Hemolymph was extracted from the third instar larvae that were immersed in the food medium (fed state). We also tested hemolymph from larvae starved for 15 hours (starved state). In both cases, the trehalose levels were significantly higher for the imp mutant compared with either control strain (Figure 2(c)). In contrast, no difference was observed among these strains for protein levels in either fed or starved condition. In order to examine the effect of imp knockdown mutation on total trehalose levels, whole-fly trehalose that is normalized by total protein levels was compared. In the starved state, the total amount of trehalose was higher than the control (Figure 2(b)). Because in our QTL analysis Niddm22 locus was identified as fasting hyperglycemic QTL, the observation of a more prominent effect on the starved state implies gene candidacy.


Use of Drosophila as an evaluation method reveals imp as a candidate gene for type 2 diabetes in rat locus Niddm22.

Kawasaki K, Yamada S, Ogata K, Saito Y, Takahama A, Yamada T, Matsumoto K, Kose H - J Diabetes Res (2015)

Expression pattern of the GFP-tagged protein trap. imp expression in third instar larva was examined using a protein trap strain ZCL0310, which expresses GFP-Imp chimeric protein in endogenous imp-expressing tissues. Bright field (a, c) and intrinsic GFP (b, d) images are shown. The GFP signal was detected in the central nervous system (CNS) (a, b), but only in a subset of neurons within the CNS. In contrast, GFP-Imp was not confirmed in the fat body (c, d). Bar, 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4363715&req=5

fig1: Expression pattern of the GFP-tagged protein trap. imp expression in third instar larva was examined using a protein trap strain ZCL0310, which expresses GFP-Imp chimeric protein in endogenous imp-expressing tissues. Bright field (a, c) and intrinsic GFP (b, d) images are shown. The GFP signal was detected in the central nervous system (CNS) (a, b), but only in a subset of neurons within the CNS. In contrast, GFP-Imp was not confirmed in the fat body (c, d). Bar, 100 μm.
Mentions: Previous studies showed that imp is expressed in the central nervous system and pole cells during embryonic development and germ cells in adults [26–28]. In order to examine the expression pattern of imp in larvae, we analyzed a protein trap strain, ZCL0310 [28]. In the third instar wandering larvae, the expression was exclusively detected in the central nervous system (CNS) (Figure 1). In contrast, imp is not expressed in other metabolically crucial tissues, including body wall muscle, fat body, gut, and oenocytes (Figure 1). Next we produced an CNS specific imp knockdown strain. The mutant had normal hatching rate and developmental growth. No morphological defect was observed. We confirmed that, in the third instar larvae, imp expression was reduced to about 20% of that of control (Figure 2(a)). Hemolymph was extracted from the third instar larvae that were immersed in the food medium (fed state). We also tested hemolymph from larvae starved for 15 hours (starved state). In both cases, the trehalose levels were significantly higher for the imp mutant compared with either control strain (Figure 2(c)). In contrast, no difference was observed among these strains for protein levels in either fed or starved condition. In order to examine the effect of imp knockdown mutation on total trehalose levels, whole-fly trehalose that is normalized by total protein levels was compared. In the starved state, the total amount of trehalose was higher than the control (Figure 2(b)). Because in our QTL analysis Niddm22 locus was identified as fasting hyperglycemic QTL, the observation of a more prominent effect on the starved state implies gene candidacy.

Bottom Line: In the mutant, the expression of two of the insulin-like peptides encoded in the fly genome, dilp2 and dilp3, were found to be downregulated.Consistent with previous reports of dilp mutants, the imp mutant flies exhibited an extension of life span; in contrast, starvation tolerance was reduced.These results further reinforce the possibility that imp is involved in sugar metabolism by modulating insulin expression.

View Article: PubMed Central - PubMed

Affiliation: Division of Natural Sciences, Department of Life Science, International Christian University, Mitaka, Tokyo 181-8585, Japan.

ABSTRACT
Type 2 diabetes (T2D) is one of the most common human diseases. QTL analysis of the diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats has identified numerous hyperglycemic loci. However, molecular characterization and/or gene identification largely remains to be elucidated due mostly to the weak genetic variances contributed by each locus. Here we utilized Drosophila melanogaster as a secondary model organism for functional evaluation of the candidate gene. We demonstrate that the tissue specific knockdown of a homologue of igf2bp2 RNA binding protein leads to increased sugar levels similar to that found in the OLETF rat. In the mutant, the expression of two of the insulin-like peptides encoded in the fly genome, dilp2 and dilp3, were found to be downregulated. Consistent with previous reports of dilp mutants, the imp mutant flies exhibited an extension of life span; in contrast, starvation tolerance was reduced. These results further reinforce the possibility that imp is involved in sugar metabolism by modulating insulin expression.

Show MeSH
Related in: MedlinePlus