Limits...
Ectopic Expression of Retrotransposon-Derived PEG11/RTL1 Contributes to the Callipyge Muscular Hypertrophy.

Xu X, Ectors F, Davis EE, Pirottin D, Cheng H, Farnir F, Hadfield T, Cockett N, Charlier C, Georges M, Takeda H - PLoS ONE (2015)

Bottom Line: The callipyge phenotype is an ovine muscular hypertrophy characterized by polar overdominance: only heterozygous +Mat/CLPGPat animals receiving the CLPG mutation from their father express the phenotype. +Mat/CLPGPat animals are characterized by postnatal, ectopic expression of Delta-like 1 homologue (DLK1) and Paternally expressed gene 11/Retrotransposon-like 1 (PEG11/RTL1) proteins in skeletal muscle.We herein describe newly generated transgenic mice that ectopically express PEG11 in skeletal muscle, and show that they also exhibit a muscular hypertrophy phenotype.Our data suggest that both DLK1 and PEG11 act together in causing the muscular hypertrophy of callipyge sheep.

View Article: PubMed Central - PubMed

Affiliation: Unit of Animal Genomics, GIGA Research Center and Faculty of Veterinary Medicine, University of Liège, 1 Avenue de l'Hôpital, Liège, Belgium.

ABSTRACT
The callipyge phenotype is an ovine muscular hypertrophy characterized by polar overdominance: only heterozygous +Mat/CLPGPat animals receiving the CLPG mutation from their father express the phenotype. +Mat/CLPGPat animals are characterized by postnatal, ectopic expression of Delta-like 1 homologue (DLK1) and Paternally expressed gene 11/Retrotransposon-like 1 (PEG11/RTL1) proteins in skeletal muscle. We showed previously in transgenic mice that ectopic expression of DLK1 alone induces a muscular hypertrophy, hence demonstrating a role for DLK1 in determining the callipyge hypertrophy. We herein describe newly generated transgenic mice that ectopically express PEG11 in skeletal muscle, and show that they also exhibit a muscular hypertrophy phenotype. Our data suggest that both DLK1 and PEG11 act together in causing the muscular hypertrophy of callipyge sheep.

No MeSH data available.


Related in: MedlinePlus

Expression analysis of the oPEG11 transgenes.A. Relative expression levels of Ccdc91 (mean ± 1.96 x standard error of the mean (s.e.m.)) measured by QRT-PCR using RNA extracted from quadriceps femoris muscles of +/+ and +/TP mice from oPEG11 line 127A. The nearly 1.5-fold reduction in +/TP animals suggests an inactivation of the Ccdc91 gene and likely explains the absence of TP/TP mice in this cross. B. Northern blotting of mRNA extracted from quadriceps femoris of +/+, +/TP, and TP/TP mice from oPEG11 lines 126 and 127A using RNA probes corresponding to the ovine PEG11 transgene (oPEG11) and mouse β–actin for control (Actb). The bands have the approximate size expected for transcripts derived from the oPEG11 transgene (4.3 Kb, arrow) in the both mouse lines plus a larger band (~7 Kb, arrow head) in line 127. C. RT-PCR (or PCR) performed using total RNA (or DNA) extracted from quadriceps femoris (or tail) of +/+, +/TP, and TP/TP mice from lines 126 and 127A using primer combinations shown in Fig 1A. The extra-bands corresponding to unspliced transcripts observed in line 127A are differentiated from the bands expected for spliced transcripts observed in lines 126 and 127A. D. Results of 5’ and 3’ RACE experiments conducted with total RNA extracted from quadriceps femoris of +/+, and +/TP mice from lines 126 and 127A. Sequencing the PCR products marked by the arrows confirmed the use of the vector-specific transcription start and polyadenylation sites (data not shown). E. Relative expression level of oPEG11 (mean ± 1.96 x s.e.m.) in line 126 measured by QRT-PCR using total RNA extracted from quadriceps femoris from animals of indicated genotypes and ages, and from tissues at 10 weeks of age. Expression levels of +/TP animals were used for references. The numbers of tested samples are shown in parentheses.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4608697&req=5

pone.0140594.g002: Expression analysis of the oPEG11 transgenes.A. Relative expression levels of Ccdc91 (mean ± 1.96 x standard error of the mean (s.e.m.)) measured by QRT-PCR using RNA extracted from quadriceps femoris muscles of +/+ and +/TP mice from oPEG11 line 127A. The nearly 1.5-fold reduction in +/TP animals suggests an inactivation of the Ccdc91 gene and likely explains the absence of TP/TP mice in this cross. B. Northern blotting of mRNA extracted from quadriceps femoris of +/+, +/TP, and TP/TP mice from oPEG11 lines 126 and 127A using RNA probes corresponding to the ovine PEG11 transgene (oPEG11) and mouse β–actin for control (Actb). The bands have the approximate size expected for transcripts derived from the oPEG11 transgene (4.3 Kb, arrow) in the both mouse lines plus a larger band (~7 Kb, arrow head) in line 127. C. RT-PCR (or PCR) performed using total RNA (or DNA) extracted from quadriceps femoris (or tail) of +/+, +/TP, and TP/TP mice from lines 126 and 127A using primer combinations shown in Fig 1A. The extra-bands corresponding to unspliced transcripts observed in line 127A are differentiated from the bands expected for spliced transcripts observed in lines 126 and 127A. D. Results of 5’ and 3’ RACE experiments conducted with total RNA extracted from quadriceps femoris of +/+, and +/TP mice from lines 126 and 127A. Sequencing the PCR products marked by the arrows confirmed the use of the vector-specific transcription start and polyadenylation sites (data not shown). E. Relative expression level of oPEG11 (mean ± 1.96 x s.e.m.) in line 126 measured by QRT-PCR using total RNA extracted from quadriceps femoris from animals of indicated genotypes and ages, and from tissues at 10 weeks of age. Expression levels of +/TP animals were used for references. The numbers of tested samples are shown in parentheses.

Mentions: Mouse line 127 was shown to harbor two distinct integration sites (127A and 127B), both comprising a similar, complex array thought to encompass multiple tandem copies of digested insert and undigested full-length vector, confirming the contamination of the insert preparation with undigested vector (Fig 1C; S2 Fig). Expression of the transgene was only observed in mice with the 127A integration, not in mice with the 127B integration (data not shown). The 127A integration site was shown to correspond to the last intron of Coiled-coil domain containing 91 (Ccdc91) on chromosome 6 (S1 Fig). Ccdc91 was shown to be highly expressed in skeletal muscle, and its expression to be reduced strongly by the transgene insertion (p = 5.6x10-5; Fig 2A).


Ectopic Expression of Retrotransposon-Derived PEG11/RTL1 Contributes to the Callipyge Muscular Hypertrophy.

Xu X, Ectors F, Davis EE, Pirottin D, Cheng H, Farnir F, Hadfield T, Cockett N, Charlier C, Georges M, Takeda H - PLoS ONE (2015)

Expression analysis of the oPEG11 transgenes.A. Relative expression levels of Ccdc91 (mean ± 1.96 x standard error of the mean (s.e.m.)) measured by QRT-PCR using RNA extracted from quadriceps femoris muscles of +/+ and +/TP mice from oPEG11 line 127A. The nearly 1.5-fold reduction in +/TP animals suggests an inactivation of the Ccdc91 gene and likely explains the absence of TP/TP mice in this cross. B. Northern blotting of mRNA extracted from quadriceps femoris of +/+, +/TP, and TP/TP mice from oPEG11 lines 126 and 127A using RNA probes corresponding to the ovine PEG11 transgene (oPEG11) and mouse β–actin for control (Actb). The bands have the approximate size expected for transcripts derived from the oPEG11 transgene (4.3 Kb, arrow) in the both mouse lines plus a larger band (~7 Kb, arrow head) in line 127. C. RT-PCR (or PCR) performed using total RNA (or DNA) extracted from quadriceps femoris (or tail) of +/+, +/TP, and TP/TP mice from lines 126 and 127A using primer combinations shown in Fig 1A. The extra-bands corresponding to unspliced transcripts observed in line 127A are differentiated from the bands expected for spliced transcripts observed in lines 126 and 127A. D. Results of 5’ and 3’ RACE experiments conducted with total RNA extracted from quadriceps femoris of +/+, and +/TP mice from lines 126 and 127A. Sequencing the PCR products marked by the arrows confirmed the use of the vector-specific transcription start and polyadenylation sites (data not shown). E. Relative expression level of oPEG11 (mean ± 1.96 x s.e.m.) in line 126 measured by QRT-PCR using total RNA extracted from quadriceps femoris from animals of indicated genotypes and ages, and from tissues at 10 weeks of age. Expression levels of +/TP animals were used for references. The numbers of tested samples are shown in parentheses.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608697&req=5

pone.0140594.g002: Expression analysis of the oPEG11 transgenes.A. Relative expression levels of Ccdc91 (mean ± 1.96 x standard error of the mean (s.e.m.)) measured by QRT-PCR using RNA extracted from quadriceps femoris muscles of +/+ and +/TP mice from oPEG11 line 127A. The nearly 1.5-fold reduction in +/TP animals suggests an inactivation of the Ccdc91 gene and likely explains the absence of TP/TP mice in this cross. B. Northern blotting of mRNA extracted from quadriceps femoris of +/+, +/TP, and TP/TP mice from oPEG11 lines 126 and 127A using RNA probes corresponding to the ovine PEG11 transgene (oPEG11) and mouse β–actin for control (Actb). The bands have the approximate size expected for transcripts derived from the oPEG11 transgene (4.3 Kb, arrow) in the both mouse lines plus a larger band (~7 Kb, arrow head) in line 127. C. RT-PCR (or PCR) performed using total RNA (or DNA) extracted from quadriceps femoris (or tail) of +/+, +/TP, and TP/TP mice from lines 126 and 127A using primer combinations shown in Fig 1A. The extra-bands corresponding to unspliced transcripts observed in line 127A are differentiated from the bands expected for spliced transcripts observed in lines 126 and 127A. D. Results of 5’ and 3’ RACE experiments conducted with total RNA extracted from quadriceps femoris of +/+, and +/TP mice from lines 126 and 127A. Sequencing the PCR products marked by the arrows confirmed the use of the vector-specific transcription start and polyadenylation sites (data not shown). E. Relative expression level of oPEG11 (mean ± 1.96 x s.e.m.) in line 126 measured by QRT-PCR using total RNA extracted from quadriceps femoris from animals of indicated genotypes and ages, and from tissues at 10 weeks of age. Expression levels of +/TP animals were used for references. The numbers of tested samples are shown in parentheses.
Mentions: Mouse line 127 was shown to harbor two distinct integration sites (127A and 127B), both comprising a similar, complex array thought to encompass multiple tandem copies of digested insert and undigested full-length vector, confirming the contamination of the insert preparation with undigested vector (Fig 1C; S2 Fig). Expression of the transgene was only observed in mice with the 127A integration, not in mice with the 127B integration (data not shown). The 127A integration site was shown to correspond to the last intron of Coiled-coil domain containing 91 (Ccdc91) on chromosome 6 (S1 Fig). Ccdc91 was shown to be highly expressed in skeletal muscle, and its expression to be reduced strongly by the transgene insertion (p = 5.6x10-5; Fig 2A).

Bottom Line: The callipyge phenotype is an ovine muscular hypertrophy characterized by polar overdominance: only heterozygous +Mat/CLPGPat animals receiving the CLPG mutation from their father express the phenotype. +Mat/CLPGPat animals are characterized by postnatal, ectopic expression of Delta-like 1 homologue (DLK1) and Paternally expressed gene 11/Retrotransposon-like 1 (PEG11/RTL1) proteins in skeletal muscle.We herein describe newly generated transgenic mice that ectopically express PEG11 in skeletal muscle, and show that they also exhibit a muscular hypertrophy phenotype.Our data suggest that both DLK1 and PEG11 act together in causing the muscular hypertrophy of callipyge sheep.

View Article: PubMed Central - PubMed

Affiliation: Unit of Animal Genomics, GIGA Research Center and Faculty of Veterinary Medicine, University of Liège, 1 Avenue de l'Hôpital, Liège, Belgium.

ABSTRACT
The callipyge phenotype is an ovine muscular hypertrophy characterized by polar overdominance: only heterozygous +Mat/CLPGPat animals receiving the CLPG mutation from their father express the phenotype. +Mat/CLPGPat animals are characterized by postnatal, ectopic expression of Delta-like 1 homologue (DLK1) and Paternally expressed gene 11/Retrotransposon-like 1 (PEG11/RTL1) proteins in skeletal muscle. We showed previously in transgenic mice that ectopic expression of DLK1 alone induces a muscular hypertrophy, hence demonstrating a role for DLK1 in determining the callipyge hypertrophy. We herein describe newly generated transgenic mice that ectopically express PEG11 in skeletal muscle, and show that they also exhibit a muscular hypertrophy phenotype. Our data suggest that both DLK1 and PEG11 act together in causing the muscular hypertrophy of callipyge sheep.

No MeSH data available.


Related in: MedlinePlus