Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis
The prion family comprises of Prion, Shadoo and Doppel proteins, encoded by Prnp, Sprn and Prnd, respectively.Expression pattern of these genes in early mouse embryogenesis was investigated using RT-PCR experiments on FVB/N mice embryosand by in situ hybridization (HIS) at 3 early developmental stag...
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| Format: | Second cycle, A2E |
| Language: | Swedish Inglés |
| Published: |
2012
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| Online Access: | https://stud.epsilon.slu.se/5096/ |
| _version_ | 1855570787565568000 |
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| author | Chachondia, Kadambari |
| author_browse | Chachondia, Kadambari |
| author_facet | Chachondia, Kadambari |
| author_sort | Chachondia, Kadambari |
| collection | Epsilon Archive for Student Projects |
| description | The prion family comprises of Prion, Shadoo and Doppel proteins, encoded by Prnp, Sprn and Prnd, respectively.Expression pattern of these genes in early mouse embryogenesis was investigated using RT-PCR experiments on FVB/N mice embryosand by in situ hybridization (HIS) at 3 early developmental stages. They were found to be expressed at all stages, in both placenta and embryo.Ubiquitous hybridization was observed for the three lociby HIS. These results are compatible with potential biological overlapping roles of these proteins during mouse embryogenesis. We also analyzed the transmission of an LS1.06 transgenic mice line, characterized by a 70% downregulationofSprngene expression.Only 5% of offspring from FVB/N LSI.06+/--Prnp+/-with FVB/N Prnp-/-crosses were of a FVB/N LSI.06+/--Prnp-/- genotype, suggestive ofa lethal-associated embryonic phenotype. However, crossing FVB/N LSI.06+/--Prnp-/-with Prnp-/- showed a Mendelian transmission rate. This suggested a physical linkage between the transgene integration site and Prnp locus. Indeed, the transgene integration site was found to be located within an intron of the mouse Api5 gene, at 37.5 Mbof thePrnp locus. It thus likely explains the observed transmission rates.
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| format | Second cycle, A2E |
| id | RepoSLU5096 |
| institution | Swedish University of Agricultural Sciences |
| language | Swedish Inglés |
| publishDate | 2012 |
| publishDateSort | 2012 |
| record_format | eprints |
| spelling | RepoSLU50962013-06-05T09:42:36Z https://stud.epsilon.slu.se/5096/ Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis Chachondia, Kadambari Animal genetics and breeding The prion family comprises of Prion, Shadoo and Doppel proteins, encoded by Prnp, Sprn and Prnd, respectively.Expression pattern of these genes in early mouse embryogenesis was investigated using RT-PCR experiments on FVB/N mice embryosand by in situ hybridization (HIS) at 3 early developmental stages. They were found to be expressed at all stages, in both placenta and embryo.Ubiquitous hybridization was observed for the three lociby HIS. These results are compatible with potential biological overlapping roles of these proteins during mouse embryogenesis. We also analyzed the transmission of an LS1.06 transgenic mice line, characterized by a 70% downregulationofSprngene expression.Only 5% of offspring from FVB/N LSI.06+/--Prnp+/-with FVB/N Prnp-/-crosses were of a FVB/N LSI.06+/--Prnp-/- genotype, suggestive ofa lethal-associated embryonic phenotype. However, crossing FVB/N LSI.06+/--Prnp-/-with Prnp-/- showed a Mendelian transmission rate. This suggested a physical linkage between the transgene integration site and Prnp locus. Indeed, the transgene integration site was found to be located within an intron of the mouse Api5 gene, at 37.5 Mbof thePrnp locus. It thus likely explains the observed transmission rates. Prions are infectious proteinaceous particles responsible for transmissible spongiform encephalopathies (TSE).The pathology and clinical disease associated with TSE are brought about by conversion of the cellular form of prion proteinPrPC, encoded by Prnp, to an infectious isoformPrPSc. Along with PrPc, two other structurally-related proteins, called Doppel and Shadoo, encoded by Prnd and Sprn, havebeen identified as its paralogs. These three genes constitute ‘the prion family’. However, the biological roles and possible biological redundancy between these proteins remain mostly enigmatic although a potential involvement during early embryogenesis was suggested. The first aim of my study was to investigate the expression pattern of the Prion gene family in early mouse embryogenesis to look for potential overlaps between them and to assess their putative role. Expression analysis of the three genes was done by RT-PCR experiments on FVB/N andFVB/N Prnp-/-embryosand by in situ hybridization (HIS) at three developmental stages. RT-PCR experiments showed lower expression pattern of Prnd and Sprncompared toPrnp. Expression was observed in all studied stages, in both placenta and embryo, except in Prnp-/- for Prnp.Ubiquitous hybridization was observed for the three lociby HIS.These results are compatible with potential biological overlapping roles and expression of these proteinsand also point out a potential role of Doppel in early embryonic stages. It would be of interest to assess the phenotypic consequences of the invalidation of Sprn and Prnd, and of the three loci. Comparative histological analysis was done between E7.5 FVB/N and FVB/N Prnp-/- mouse embryos. The phenotype was similar except for the observation of hemorrhagic foci in front of ectoplacental cone in FVB/N.This could be due to the involvement of PrPC in biological pathways like angiogenesis, inflammation and cell mobility. The second aim of my study was the analysis ofthe LSI.06 transgenic line and of thetransmission of its transgene onto Prnp-/-genetic backgrounds. This line expresses ubiquitously a ShRNA against Sprn and has 70% downregulation of Sprn expression as observed in the adult brain. Transmission studies weredone by crossing FVB/N LSI.06+/--Prnp+/- with Prnp-/- mice. FVB/N LSI.06+/--Prnp-/-and LSI.06-/--Prnp+/-offsprings were less than 5% and 16%, respectively. LSI.06+/--Prnp+/- mice were also crossed with C57/129/Sv Prnp-/-animals to analyze the incidence of the genetic background associated with Prnp-/-. There was again a non-Mendelian transmission with deficiency in LSI.06+/--Prnp-/- and LSI.06-/--Prnp+/- offsprings.FVB/N LSI.06+/--Prnp-/-were crossed with FVB/N Prnp-/- mice to analyze secondary transmission rate of the LSI.O6 transgene. It showed a Mendelian [3] transmission rate. This data suggested a physical linkage between theLSI.06 transgene integration site and thePrnp locus rather than a lethal-associated embryonic phenotype. However, assessment of the fecundity of such mice, including growth rate and robustness of pups might give us a possible clue for what are the further effects of such introgression.The transgene integration site was cloned and found to be located within the last intron of the ORF of the mouse Api5 (apoptosis inhibitor 5) gene,at 37.5 Mbof thePrnp locus. It thus likely explains the observed transmission rates. 2012-12-04 Second cycle, A2E NonPeerReviewed application/pdf sv https://stud.epsilon.slu.se/5096/13/kadambari_ch_121205.pdf Chachondia, Kadambari, 2012. Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis. Second cycle, A2E. Uppsala: (VH) > Dept. of Animal Breeding and Genetics (until 231231) <https://stud.epsilon.slu.se/view/divisions/OID-670.html> urn:nbn:se:slu:epsilon-s-1897 eng |
| spellingShingle | Animal genetics and breeding Chachondia, Kadambari Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title | Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title_full | Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title_fullStr | Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title_full_unstemmed | Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title_short | Analysis of the biological role of prion-related Shadoo protein on early mouse embryogenesis |
| title_sort | analysis of the biological role of prion-related shadoo protein on early mouse embryogenesis |
| topic | Animal genetics and breeding |
| url | https://stud.epsilon.slu.se/5096/ https://stud.epsilon.slu.se/5096/ |