SRSF2 P95H MUTATION RESULTS IN IMPAIRED STEM CELL REPOPULATION AND COMPROMISED HEMATOPOIETIC DIFFERENTIATION IN MICE
(Abstract release date: 05/21/15)
EHA Library. Kon A. 06/12/15; 103217; S134
Disclosure(s): Kyoto UniversityDept. Pathology and Tumor Biology
Dr. Ayana Kon
Contributions
Contributions
Abstract
Abstract: S134
Type: Oral Presentation + travel grant
Presentation during EHA20: From 12.06.2015 12:15 to 12.06.2015 12:30
Location: Room Lehar 3 + 4
Background
Recent genetic studies have revealed frequent and specific pathway mutations involving multiple components of the RNA splicing machinery in myelodysplasia. Among these, SRSF2 mutations are more prevalent in CMML subtype and are associated with poor prognosis. Mutations showed a prominent hotspot involving proline 95, causing either P95H, P95L, or P95R conversion. The molecular mechanism by which SRSF2 mutations lead to myelodysplasia remains largely unknown.
Aims
This study aimed to clarify the role of SRSF2 mutations in the development of myelodysplasia through the analysis of Srsf2 P95H conditional knock-in mice.
Methods
We first generated a heterozygous conditional knock-in mouse model of Srsf2 P95H mutation and crossed them with Vav1-Cre transgenic mice. We then performed detailed analysis of hematological phenotype of Srsf2 P95H knock-in mice and also evaluated their reconstitution ability in competitive transplantation experiments.
Results
Heterozygous Srsf2 P95H conditional knock-in mice exhibited no significant change in total peripheral blood (PB) counts compared to wild-type mice at 8-15 weeks after birth. Bone marrow (BM) cellularity and spleen weight showed no obvious difference between Srsf2 P95H and wild-type mice. Analysis of hematopoietic stem and progenitor cells fractions showed significant decrease of the frequency of HSCs in Srsf2 P95H mice compared to wild-type mice. On the other hand, there were no significant differences in the number of more differentiated progenitor cells including multipotent progenitor (MPP) cell fractions, myeloid progenitors (MEPs, CMPs, and GMPs) and lymphoid progenitors between Srsf2 P95H and wild-type mice. We next performed noncompetitive transplantation experiments to assess the cell intrinsic effects of Srsf2 P95H mutations. 3 months after transplantation, recipient mice transplanted with Srsf2 P95H BM cells showed significant leukopenia due to impaired lymphopoiesis and anemia. Flow cytometrical analysis revealed decreased numbers of HSCs and MPPs fractions, whereas there were no significant changes in MEPs, CMPs, and GMPs in BM. The frequency of erythroid progenitor populations was significantly reduced in Srsf2 P95H mice in BM and spleens. The population of B cell lineage also decreased at as early as pre-pro B cell stages. These observations suggested that SRSF2 P95H mutations lead to impaired stem cell functions and ineffective hematopoiesis, resembling the phenotype of MDS. Subsequently, we assessed the reconstitution capacity of whole BM cells from SRSF2 mutant mice in competitive transplantation experiments. The donor chimerism of Srsf2 P95H-derived cells in PB was significantly lower than that of wild-type cells. At 4 months post transplantation, chimerism of Srsf2 P95H-derived cells was remarkably lower than that of wild-type cells in the fractions of HSCs, MPPs, CMPs, MEPs, GMPs and CLPs. Furthermore, the reduced donor chimerism for Srsf2 P95H mutants was recapitulated in secondary transplantation experiments. Finally, we have not observed overt MDS phenotypes in none of these Srsf2-mutant mice or transplantation models.
Summary
Our results demonstrated that heterozygous P95H mutation of Srsf2 lead to deregulation of hematopoietic stem cells that was evident from reduced competitive repopulation in lethally irradiated mice and impaired hematopoietic differentiation. SRSF2 mutation by itself does not seem to be sufficient to develop MDS but to require additional genetic and/or epigenetic events for overt MDS phenotype.
Keyword(s): Mouse model, Myelodysplasia, Somatic mutation
Session topic: Biology in MDS
Type: Oral Presentation + travel grant
Presentation during EHA20: From 12.06.2015 12:15 to 12.06.2015 12:30
Location: Room Lehar 3 + 4
Background
Recent genetic studies have revealed frequent and specific pathway mutations involving multiple components of the RNA splicing machinery in myelodysplasia. Among these, SRSF2 mutations are more prevalent in CMML subtype and are associated with poor prognosis. Mutations showed a prominent hotspot involving proline 95, causing either P95H, P95L, or P95R conversion. The molecular mechanism by which SRSF2 mutations lead to myelodysplasia remains largely unknown.
Aims
This study aimed to clarify the role of SRSF2 mutations in the development of myelodysplasia through the analysis of Srsf2 P95H conditional knock-in mice.
Methods
We first generated a heterozygous conditional knock-in mouse model of Srsf2 P95H mutation and crossed them with Vav1-Cre transgenic mice. We then performed detailed analysis of hematological phenotype of Srsf2 P95H knock-in mice and also evaluated their reconstitution ability in competitive transplantation experiments.
Results
Heterozygous Srsf2 P95H conditional knock-in mice exhibited no significant change in total peripheral blood (PB) counts compared to wild-type mice at 8-15 weeks after birth. Bone marrow (BM) cellularity and spleen weight showed no obvious difference between Srsf2 P95H and wild-type mice. Analysis of hematopoietic stem and progenitor cells fractions showed significant decrease of the frequency of HSCs in Srsf2 P95H mice compared to wild-type mice. On the other hand, there were no significant differences in the number of more differentiated progenitor cells including multipotent progenitor (MPP) cell fractions, myeloid progenitors (MEPs, CMPs, and GMPs) and lymphoid progenitors between Srsf2 P95H and wild-type mice. We next performed noncompetitive transplantation experiments to assess the cell intrinsic effects of Srsf2 P95H mutations. 3 months after transplantation, recipient mice transplanted with Srsf2 P95H BM cells showed significant leukopenia due to impaired lymphopoiesis and anemia. Flow cytometrical analysis revealed decreased numbers of HSCs and MPPs fractions, whereas there were no significant changes in MEPs, CMPs, and GMPs in BM. The frequency of erythroid progenitor populations was significantly reduced in Srsf2 P95H mice in BM and spleens. The population of B cell lineage also decreased at as early as pre-pro B cell stages. These observations suggested that SRSF2 P95H mutations lead to impaired stem cell functions and ineffective hematopoiesis, resembling the phenotype of MDS. Subsequently, we assessed the reconstitution capacity of whole BM cells from SRSF2 mutant mice in competitive transplantation experiments. The donor chimerism of Srsf2 P95H-derived cells in PB was significantly lower than that of wild-type cells. At 4 months post transplantation, chimerism of Srsf2 P95H-derived cells was remarkably lower than that of wild-type cells in the fractions of HSCs, MPPs, CMPs, MEPs, GMPs and CLPs. Furthermore, the reduced donor chimerism for Srsf2 P95H mutants was recapitulated in secondary transplantation experiments. Finally, we have not observed overt MDS phenotypes in none of these Srsf2-mutant mice or transplantation models.
Summary
Our results demonstrated that heterozygous P95H mutation of Srsf2 lead to deregulation of hematopoietic stem cells that was evident from reduced competitive repopulation in lethally irradiated mice and impaired hematopoietic differentiation. SRSF2 mutation by itself does not seem to be sufficient to develop MDS but to require additional genetic and/or epigenetic events for overt MDS phenotype.
Keyword(s): Mouse model, Myelodysplasia, Somatic mutation
Session topic: Biology in MDS
Abstract: S134
Type: Oral Presentation + travel grant
Presentation during EHA20: From 12.06.2015 12:15 to 12.06.2015 12:30
Location: Room Lehar 3 + 4
Background
Recent genetic studies have revealed frequent and specific pathway mutations involving multiple components of the RNA splicing machinery in myelodysplasia. Among these, SRSF2 mutations are more prevalent in CMML subtype and are associated with poor prognosis. Mutations showed a prominent hotspot involving proline 95, causing either P95H, P95L, or P95R conversion. The molecular mechanism by which SRSF2 mutations lead to myelodysplasia remains largely unknown.
Aims
This study aimed to clarify the role of SRSF2 mutations in the development of myelodysplasia through the analysis of Srsf2 P95H conditional knock-in mice.
Methods
We first generated a heterozygous conditional knock-in mouse model of Srsf2 P95H mutation and crossed them with Vav1-Cre transgenic mice. We then performed detailed analysis of hematological phenotype of Srsf2 P95H knock-in mice and also evaluated their reconstitution ability in competitive transplantation experiments.
Results
Heterozygous Srsf2 P95H conditional knock-in mice exhibited no significant change in total peripheral blood (PB) counts compared to wild-type mice at 8-15 weeks after birth. Bone marrow (BM) cellularity and spleen weight showed no obvious difference between Srsf2 P95H and wild-type mice. Analysis of hematopoietic stem and progenitor cells fractions showed significant decrease of the frequency of HSCs in Srsf2 P95H mice compared to wild-type mice. On the other hand, there were no significant differences in the number of more differentiated progenitor cells including multipotent progenitor (MPP) cell fractions, myeloid progenitors (MEPs, CMPs, and GMPs) and lymphoid progenitors between Srsf2 P95H and wild-type mice. We next performed noncompetitive transplantation experiments to assess the cell intrinsic effects of Srsf2 P95H mutations. 3 months after transplantation, recipient mice transplanted with Srsf2 P95H BM cells showed significant leukopenia due to impaired lymphopoiesis and anemia. Flow cytometrical analysis revealed decreased numbers of HSCs and MPPs fractions, whereas there were no significant changes in MEPs, CMPs, and GMPs in BM. The frequency of erythroid progenitor populations was significantly reduced in Srsf2 P95H mice in BM and spleens. The population of B cell lineage also decreased at as early as pre-pro B cell stages. These observations suggested that SRSF2 P95H mutations lead to impaired stem cell functions and ineffective hematopoiesis, resembling the phenotype of MDS. Subsequently, we assessed the reconstitution capacity of whole BM cells from SRSF2 mutant mice in competitive transplantation experiments. The donor chimerism of Srsf2 P95H-derived cells in PB was significantly lower than that of wild-type cells. At 4 months post transplantation, chimerism of Srsf2 P95H-derived cells was remarkably lower than that of wild-type cells in the fractions of HSCs, MPPs, CMPs, MEPs, GMPs and CLPs. Furthermore, the reduced donor chimerism for Srsf2 P95H mutants was recapitulated in secondary transplantation experiments. Finally, we have not observed overt MDS phenotypes in none of these Srsf2-mutant mice or transplantation models.
Summary
Our results demonstrated that heterozygous P95H mutation of Srsf2 lead to deregulation of hematopoietic stem cells that was evident from reduced competitive repopulation in lethally irradiated mice and impaired hematopoietic differentiation. SRSF2 mutation by itself does not seem to be sufficient to develop MDS but to require additional genetic and/or epigenetic events for overt MDS phenotype.
Keyword(s): Mouse model, Myelodysplasia, Somatic mutation
Session topic: Biology in MDS
Type: Oral Presentation + travel grant
Presentation during EHA20: From 12.06.2015 12:15 to 12.06.2015 12:30
Location: Room Lehar 3 + 4
Background
Recent genetic studies have revealed frequent and specific pathway mutations involving multiple components of the RNA splicing machinery in myelodysplasia. Among these, SRSF2 mutations are more prevalent in CMML subtype and are associated with poor prognosis. Mutations showed a prominent hotspot involving proline 95, causing either P95H, P95L, or P95R conversion. The molecular mechanism by which SRSF2 mutations lead to myelodysplasia remains largely unknown.
Aims
This study aimed to clarify the role of SRSF2 mutations in the development of myelodysplasia through the analysis of Srsf2 P95H conditional knock-in mice.
Methods
We first generated a heterozygous conditional knock-in mouse model of Srsf2 P95H mutation and crossed them with Vav1-Cre transgenic mice. We then performed detailed analysis of hematological phenotype of Srsf2 P95H knock-in mice and also evaluated their reconstitution ability in competitive transplantation experiments.
Results
Heterozygous Srsf2 P95H conditional knock-in mice exhibited no significant change in total peripheral blood (PB) counts compared to wild-type mice at 8-15 weeks after birth. Bone marrow (BM) cellularity and spleen weight showed no obvious difference between Srsf2 P95H and wild-type mice. Analysis of hematopoietic stem and progenitor cells fractions showed significant decrease of the frequency of HSCs in Srsf2 P95H mice compared to wild-type mice. On the other hand, there were no significant differences in the number of more differentiated progenitor cells including multipotent progenitor (MPP) cell fractions, myeloid progenitors (MEPs, CMPs, and GMPs) and lymphoid progenitors between Srsf2 P95H and wild-type mice. We next performed noncompetitive transplantation experiments to assess the cell intrinsic effects of Srsf2 P95H mutations. 3 months after transplantation, recipient mice transplanted with Srsf2 P95H BM cells showed significant leukopenia due to impaired lymphopoiesis and anemia. Flow cytometrical analysis revealed decreased numbers of HSCs and MPPs fractions, whereas there were no significant changes in MEPs, CMPs, and GMPs in BM. The frequency of erythroid progenitor populations was significantly reduced in Srsf2 P95H mice in BM and spleens. The population of B cell lineage also decreased at as early as pre-pro B cell stages. These observations suggested that SRSF2 P95H mutations lead to impaired stem cell functions and ineffective hematopoiesis, resembling the phenotype of MDS. Subsequently, we assessed the reconstitution capacity of whole BM cells from SRSF2 mutant mice in competitive transplantation experiments. The donor chimerism of Srsf2 P95H-derived cells in PB was significantly lower than that of wild-type cells. At 4 months post transplantation, chimerism of Srsf2 P95H-derived cells was remarkably lower than that of wild-type cells in the fractions of HSCs, MPPs, CMPs, MEPs, GMPs and CLPs. Furthermore, the reduced donor chimerism for Srsf2 P95H mutants was recapitulated in secondary transplantation experiments. Finally, we have not observed overt MDS phenotypes in none of these Srsf2-mutant mice or transplantation models.
Summary
Our results demonstrated that heterozygous P95H mutation of Srsf2 lead to deregulation of hematopoietic stem cells that was evident from reduced competitive repopulation in lethally irradiated mice and impaired hematopoietic differentiation. SRSF2 mutation by itself does not seem to be sufficient to develop MDS but to require additional genetic and/or epigenetic events for overt MDS phenotype.
Keyword(s): Mouse model, Myelodysplasia, Somatic mutation
Session topic: Biology in MDS
{{ help_message }}
{{filter}}