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Maternal influence of prolyl endopeptidase on fat mass of adult progeny.

Warden CH, Fisler JS, Espinal G, Graham J, Havel PJ, Perroud B - Int J Obes (Lond) (2009)

Bottom Line: There were no genotype effects on BW or AI in males or females on either diet.Experiment 2-In contrast, reciprocal crosses of heterozygous males or females with WT B6 revealed highly significant parent of origin effects on all traits except body length.Heterozygosity for PREP GT results in highly significant maternal effects, whereas homozygosity for the PREP(gt/gt) mutation has a much more limited direct effect.

View Article: PubMed Central - PubMed

Affiliation: Rowe Program in Genetics, University of California, Davis, CA 95616, USA. chwarden@ucdavis.edu

ABSTRACT

Background: Maternal genotype has lifetime effects on progeny, but few specific genes, and no proteases, are known to underlie maternal effects. Prolyl endopeptidase (PREP) is a serine protease with putative substrates that regulate appetite or milk production.

Objective: To test effects of PREP on obesity phenotypes in mice.

Design: Mice with a gene trap (GT) of PREP (PREP(gt/gt)) on the C57BL/6J (B6) background were generated. Minimal PREP protein was detected by western blot. In Experiment 1, direct effects of PREP were measured in littermate mice derived from intercrosses of heterozygotes (PREP(WT/gt)). In Experiment 2, maternal effects of PREP were measured in reciprocal crosses of heterozygous (PREP(WT/gt)) and wild-type (WT) (PREP(WT/WT)) males and females. DIETS: Mice were fed either low-fat (LF, Experiments 1 and 2) or high-fat (HF, Experiment 1) defined diets.

Measurements: Adiposity index (AI) was calculated from body weight (BW) and weights of four fat depots measured in 120-day-old mice. Fasting plasma glucose, insulin and leptin were measured. In vivo plasma alpha-MSH levels were measured by targeted quantitative peptidomics.

Results: Experiment 1-In intercross mice, there were significant diet effects, but few genotype effects. There were no genotype effects on BW or AI in males or females on either diet. Experiment 2-In contrast, reciprocal crosses of heterozygous males or females with WT B6 revealed highly significant parent of origin effects on all traits except body length. Progeny (WT and heterozygous genotypes and both sexes) born to female PREP(WT/gt) heterozygotes had fat pads that weighed as much as -twofold more at 120 days old than progeny born to male heterozygotes.

Conclusion: Heterozygosity for PREP GT results in highly significant maternal effects, whereas homozygosity for the PREP(gt/gt) mutation has a much more limited direct effect.

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Related in: MedlinePlus

Western blot of PREP protein in tissues of PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either low-fat or high-fat diets as well as recombinant PREP protein. See text for methods. The minimal PREP protein in PREPgt/gt mice indicates that this mutation is a PREP hypomorph rather than a complete deletion.
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Figure 1: Western blot of PREP protein in tissues of PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either low-fat or high-fat diets as well as recombinant PREP protein. See text for methods. The minimal PREP protein in PREPgt/gt mice indicates that this mutation is a PREP hypomorph rather than a complete deletion.

Mentions: Surveys of mRNA abundance, both BioGPS (https://biogps.gnf.org/), and MPSS GEO (http://www.ncbi.nlm.nih.gov/geo/info/mouse-trans.html) (19), reveal that PREP is expressed in several tissues. Deficiency of PREP protein in gene-trap mice was confirmed by performing a Western blot of brain and kidney proteins from PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either the LF or HF diet as shown in Figure 1A and lung from mice fed only the LF diet in Figure 1B. Additional Western blots comparing brain, muscle, ovary, uterus, lung and heart in PREPWT/WT and PREPgt/gt mice fed only the LF diet are in Figure 1C. PREP was identified by including recombinant PREP on the Western blot. The sample genotypes were verified by PCR. The antibody to PREP was made to a synthetic peptide based on the amino terminal end of human Prolyl Endopeptidase. PREP protein was present in PREPWT/WT mice, reduced in PREPWT/gt , and almost absent in PREPgt/gt mice (Figure 1). There was no detectable effect of diet on the amount of PREP protein in kidney and brain. The amount of PREP in PREPgt/gt homozygotes varied from undetectable in gastrocnemius muscle to easily visible in brain. Detectable PREP protein in PREPgt/gt mice results from alternative splicing to remove the ß-gal containing “gene-trap” exon. Our data demonstrate that efficiency of alternative splicing of PREP varies between tissues, but in all tissues PREPgt/gt exhibit reduced protein.


Maternal influence of prolyl endopeptidase on fat mass of adult progeny.

Warden CH, Fisler JS, Espinal G, Graham J, Havel PJ, Perroud B - Int J Obes (Lond) (2009)

Western blot of PREP protein in tissues of PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either low-fat or high-fat diets as well as recombinant PREP protein. See text for methods. The minimal PREP protein in PREPgt/gt mice indicates that this mutation is a PREP hypomorph rather than a complete deletion.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2743752&req=5

Figure 1: Western blot of PREP protein in tissues of PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either low-fat or high-fat diets as well as recombinant PREP protein. See text for methods. The minimal PREP protein in PREPgt/gt mice indicates that this mutation is a PREP hypomorph rather than a complete deletion.
Mentions: Surveys of mRNA abundance, both BioGPS (https://biogps.gnf.org/), and MPSS GEO (http://www.ncbi.nlm.nih.gov/geo/info/mouse-trans.html) (19), reveal that PREP is expressed in several tissues. Deficiency of PREP protein in gene-trap mice was confirmed by performing a Western blot of brain and kidney proteins from PREPWT/WT, PREPWT/gt and PREPgt/gt mice fed either the LF or HF diet as shown in Figure 1A and lung from mice fed only the LF diet in Figure 1B. Additional Western blots comparing brain, muscle, ovary, uterus, lung and heart in PREPWT/WT and PREPgt/gt mice fed only the LF diet are in Figure 1C. PREP was identified by including recombinant PREP on the Western blot. The sample genotypes were verified by PCR. The antibody to PREP was made to a synthetic peptide based on the amino terminal end of human Prolyl Endopeptidase. PREP protein was present in PREPWT/WT mice, reduced in PREPWT/gt , and almost absent in PREPgt/gt mice (Figure 1). There was no detectable effect of diet on the amount of PREP protein in kidney and brain. The amount of PREP in PREPgt/gt homozygotes varied from undetectable in gastrocnemius muscle to easily visible in brain. Detectable PREP protein in PREPgt/gt mice results from alternative splicing to remove the ß-gal containing “gene-trap” exon. Our data demonstrate that efficiency of alternative splicing of PREP varies between tissues, but in all tissues PREPgt/gt exhibit reduced protein.

Bottom Line: There were no genotype effects on BW or AI in males or females on either diet.Experiment 2-In contrast, reciprocal crosses of heterozygous males or females with WT B6 revealed highly significant parent of origin effects on all traits except body length.Heterozygosity for PREP GT results in highly significant maternal effects, whereas homozygosity for the PREP(gt/gt) mutation has a much more limited direct effect.

View Article: PubMed Central - PubMed

Affiliation: Rowe Program in Genetics, University of California, Davis, CA 95616, USA. chwarden@ucdavis.edu

ABSTRACT

Background: Maternal genotype has lifetime effects on progeny, but few specific genes, and no proteases, are known to underlie maternal effects. Prolyl endopeptidase (PREP) is a serine protease with putative substrates that regulate appetite or milk production.

Objective: To test effects of PREP on obesity phenotypes in mice.

Design: Mice with a gene trap (GT) of PREP (PREP(gt/gt)) on the C57BL/6J (B6) background were generated. Minimal PREP protein was detected by western blot. In Experiment 1, direct effects of PREP were measured in littermate mice derived from intercrosses of heterozygotes (PREP(WT/gt)). In Experiment 2, maternal effects of PREP were measured in reciprocal crosses of heterozygous (PREP(WT/gt)) and wild-type (WT) (PREP(WT/WT)) males and females. DIETS: Mice were fed either low-fat (LF, Experiments 1 and 2) or high-fat (HF, Experiment 1) defined diets.

Measurements: Adiposity index (AI) was calculated from body weight (BW) and weights of four fat depots measured in 120-day-old mice. Fasting plasma glucose, insulin and leptin were measured. In vivo plasma alpha-MSH levels were measured by targeted quantitative peptidomics.

Results: Experiment 1-In intercross mice, there were significant diet effects, but few genotype effects. There were no genotype effects on BW or AI in males or females on either diet. Experiment 2-In contrast, reciprocal crosses of heterozygous males or females with WT B6 revealed highly significant parent of origin effects on all traits except body length. Progeny (WT and heterozygous genotypes and both sexes) born to female PREP(WT/gt) heterozygotes had fat pads that weighed as much as -twofold more at 120 days old than progeny born to male heterozygotes.

Conclusion: Heterozygosity for PREP GT results in highly significant maternal effects, whereas homozygosity for the PREP(gt/gt) mutation has a much more limited direct effect.

Show MeSH
Related in: MedlinePlus