In Vitro Validation of DNA-Diffusion and Control Sequences within the AXIN2 Locus [nanopore_plasmid]
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ABSTRACT:
PROVIDER: PRJNA1124122 | ENA |
REPOSITORIES: ENA
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Project description:DNA-Diffusion is a novel generative approach leveraging diffusion probabilistic models for the design of cell type-specific DNA regulatory sequences. To evaluate the capacity of DNA-Diffusion sequences to endogenously alter AXIN2 transcription, we employed a novel MPRA-like system that utilizes CRE-recombinase mediated cassette exchange and long-read sequencing to measure the gene’s transcriptional output in response to enhancer sequences several kilobases away. We evaluated a total of 100 sequences using the AXIN2 endogenous replacement experiment using the following sequence groups: GM12878 DNA-Diffusion, GM12878 Positive Controls, GM12878 Negative Controls, K562 DNA-Diffusion, HepG2 DNA-Diffusion, and shuffled GM12878 DNA-Diffusion. Our findings demonstrate the potential of DNA-Diffusion to design sequences with therapeutic potential, showing their effectiveness in an endogenous setting.
2025-07-13 | GSE269912 | GEO
Project description:DNA-Diffusion is a novel generative approach leveraging diffusion probabilistic models for the design of cell type-specific DNA regulatory sequences. To evaluate the capacity of DNA-Diffusion sequences to endogenously alter AXIN2 transcription, we employed a novel MPRA-like system that utilizes CRE-recombinase mediated cassette exchange and long-read sequencing to measure the gene’s transcriptional output in response to enhancer sequences several kilobases away. We evaluated a total of 100 sequences using the AXIN2 endogenous replacement experiment using the following sequence groups: GM12878 DNA-Diffusion, GM12878 Positive Controls, GM12878 Negative Controls, K562 DNA-Diffusion, HepG2 DNA-Diffusion, and shuffled GM12878 DNA-Diffusion. Our findings demonstrate the potential of DNA-Diffusion to design sequences with therapeutic potential, showing their effectiveness in an endogenous setting.
2025-07-13 | GSE269915 | GEO
Project description:DNA-Diffusion is a novel generative approach leveraging diffusion probabilistic models for the design of cell type-specific DNA regulatory sequences. To evaluate the capacity of DNA-Diffusion sequences to endogenously alter AXIN2 transcription, we employed a novel MPRA-like system that utilizes CRE-recombinase mediated cassette exchange and long-read sequencing to measure the gene’s transcriptional output in response to enhancer sequences several kilobases away. We evaluated a total of 100 sequences using the AXIN2 endogenous replacement experiment using the following sequence groups: GM12878 DNA-Diffusion, GM12878 Positive Controls, GM12878 Negative Controls, K562 DNA-Diffusion, HepG2 DNA-Diffusion, and shuffled GM12878 DNA-Diffusion. Our findings demonstrate the potential of DNA-Diffusion to design sequences with therapeutic potential, showing their effectiveness in an endogenous setting.
2025-07-13 | GSE269921 | GEO
Project description:In Vitro Validation of DNA-Diffusion and Control Sequences within the AXIN2 Locus [mRNA]
| PRJNA1124111 | ENA
Project description:In Vitro Validation of DNA-Diffusion and Control Sequences within the AXIN2 Locus [gRNA]
| PRJNA1124115 | ENA
Project description:Runx2 and Axin2 regulate skeletal development. We recently determined that Axin2 and Runx2 molecularly interact in differentiating osteoblasts to regulate intramembranous bone formation, but the relationship between these factors in endochondral bone formation was unresolved. To address this, we examined the effects of Axin2 deficiency on the cleidocranial dysplasia (CCD) phenotype of Runx2+/- mice, focusing on skeletal defects attributed to improper endochondral bone formation. Axin2 deficiency unexpectedly exacerbated calvarial components of the CCD phenotype in the Runx2+/- mice; the endocranial layer of the frontal suture, which develops by endochondral bone formation, failed to mineralize in the Axin2-/-:Runx2+/-mice, resulting in a cartilaginous, fibrotic and larger fontanel than observed in Runx2+/- mice. Transcripts associated with cartilage development (e.g., Acan, miR140) were expressed at higher levels, whereas blood vessel morphogenesis transcripts (e.g., Slit2) were suppressed in Axin2-/-:Runx2+/-calvaria. Cartilage maturation was impaired, as primary chondrocytes from double mutant mice demonstrated delayed differentiation and produced less calcified matrix in vitro. The genetic dominance of Runx2 was also reflected during endochondral fracture repair, as both Runx2+/- and double mutant Axin2-/-:Runx2+/- mice had enlarged fracture calluses at early stages of healing. However, by the end stages of fracture healing, double mutant animals diverged from the Runx2+/- mice, showing smaller calluses and increased torsional strength indicative of more rapid end stage bone formation as seen in the Axin2-/- mice. Taken together, our data demonstrate a dominant role for Runx2 in chondrocyte maturation, but implicate Axin2 as an important modulator of the terminal stages of endochondral bone formation. 4 mice per genotype X 4 genotypes: wildtype (WT), Runx2+/- (R-Het), Axin2-/- (A-KO), Axin2-/-:Runx2+/- (A-KO:R-Het). Total = 16 samples
2014-07-04 | E-GEOD-59079 | biostudies-arrayexpress
Project description:Runx2 and Axin2 regulate skeletal development. We recently determined that Axin2 and Runx2 molecularly interact in differentiating osteoblasts to regulate intramembranous bone formation, but the relationship between these factors in endochondral bone formation was unresolved. To address this, we examined the effects of Axin2 deficiency on the cleidocranial dysplasia (CCD) phenotype of Runx2+/-M-BM- mice, focusing on skeletal defects attributed to improper endochondral bone formation. Axin2 deficiency unexpectedly exacerbated calvarial components of the CCD phenotype in the Runx2+/-M-BM- mice; the endocranial layer of the frontal suture, which develops by endochondral bone formation, failed to mineralize in the Axin2-/-:Runx2+/-mice, resulting in a cartilaginous, fibrotic and larger fontanel than observed in Runx2+/-M-BM- mice. Transcripts associated with cartilage development (e.g., Acan, miR140) were expressed at higher levels, whereas blood vessel morphogenesis transcripts (e.g., Slit2) were suppressed in Axin2-/-:Runx2+/-calvaria. Cartilage maturation was impaired, as primary chondrocytes from double mutant mice demonstrated delayed differentiation and produced less calcified matrix in vitro. The genetic dominance of Runx2 was also reflected during endochondral fracture repair, as both Runx2+/-M-BM- and double mutant Axin2-/-:Runx2+/-M-BM- mice had enlarged fracture calluses at early stages of healing. However, by the end stages of fracture healing, double mutant animals diverged from the Runx2+/-M-BM- mice, showing smaller calluses and increased torsional strength indicative of more rapid end stage bone formation as seen in the Axin2-/-M-BM- mice. Taken together, our data demonstrate a dominant role for Runx2 in chondrocyte maturation, but implicate Axin2 as an important modulator of the terminal stages of endochondral bone formation. 4 mice per genotype X 4 genotypes: wildtype (WT), Runx2+/- (R-Het), Axin2-/- (A-KO), Axin2-/-:Runx2+/- (A-KO:R-Het). Total = 16 samples
2014-07-04 | E-GEOD-59081 | biostudies-arrayexpress
Project description:Runx2 and Axin2 regulate skeletal development. We recently determined that Axin2 and Runx2 molecularly interact in differentiating osteoblasts to regulate intramembranous bone formation, but the relationship between these factors in endochondral bone formation was unresolved. To address this, we examined the effects of Axin2 deficiency on the cleidocranial dysplasia (CCD) phenotype of Runx2+/- mice, focusing on skeletal defects attributed to improper endochondral bone formation. Axin2 deficiency unexpectedly exacerbated calvarial components of the CCD phenotype in the Runx2+/- mice; the endocranial layer of the frontal suture, which develops by endochondral bone formation, failed to mineralize in the Axin2-/-:Runx2+/-mice, resulting in a cartilaginous, fibrotic and larger fontanel than observed in Runx2+/- mice. Transcripts associated with cartilage development (e.g., Acan, miR140) were expressed at higher levels, whereas blood vessel morphogenesis transcripts (e.g., Slit2) were suppressed in Axin2-/-:Runx2+/-calvaria. Cartilage maturation was impaired, as primary chondrocytes from double mutant mice demonstrated delayed differentiation and produced less calcified matrix in vitro. The genetic dominance of Runx2 was also reflected during endochondral fracture repair, as both Runx2+/- and double mutant Axin2-/-:Runx2+/- mice had enlarged fracture calluses at early stages of healing. However, by the end stages of fracture healing, double mutant animals diverged from the Runx2+/- mice, showing smaller calluses and increased torsional strength indicative of more rapid end stage bone formation as seen in the Axin2-/- mice. Taken together, our data demonstrate a dominant role for Runx2 in chondrocyte maturation, but implicate Axin2 as an important modulator of the terminal stages of endochondral bone formation.
2014-07-04 | GSE59081 | GEO
Project description:Runx2 and Axin2 regulate skeletal development. We recently determined that Axin2 and Runx2 molecularly interact in differentiating osteoblasts to regulate intramembranous bone formation, but the relationship between these factors in endochondral bone formation was unresolved. To address this, we examined the effects of Axin2 deficiency on the cleidocranial dysplasia (CCD) phenotype of Runx2+/- mice, focusing on skeletal defects attributed to improper endochondral bone formation. Axin2 deficiency unexpectedly exacerbated calvarial components of the CCD phenotype in the Runx2+/- mice; the endocranial layer of the frontal suture, which develops by endochondral bone formation, failed to mineralize in the Axin2-/-:Runx2+/-mice, resulting in a cartilaginous, fibrotic and larger fontanel than observed in Runx2+/- mice. Transcripts associated with cartilage development (e.g., Acan, miR140) were expressed at higher levels, whereas blood vessel morphogenesis transcripts (e.g., Slit2) were suppressed in Axin2-/-:Runx2+/-calvaria. Cartilage maturation was impaired, as primary chondrocytes from double mutant mice demonstrated delayed differentiation and produced less calcified matrix in vitro. The genetic dominance of Runx2 was also reflected during endochondral fracture repair, as both Runx2+/- and double mutant Axin2-/-:Runx2+/- mice had enlarged fracture calluses at early stages of healing. However, by the end stages of fracture healing, double mutant animals diverged from the Runx2+/- mice, showing smaller calluses and increased torsional strength indicative of more rapid end stage bone formation as seen in the Axin2-/- mice. Taken together, our data demonstrate a dominant role for Runx2 in chondrocyte maturation, but implicate Axin2 as an important modulator of the terminal stages of endochondral bone formation.
2014-07-04 | GSE59079 | GEO
Project description:A tendon’s ordered extracellular matrix (ECM) is integral for transmitting force and highly prone to injury. Whether and how tendon cells, or tenocytes, embedded within this dense ECM mobilize and contribute to healing is unknown. Here, we identify a specialized Axin2+ tenocyte population in mouse and human tendons that remains latent in homeostasis yet serves as a major source of tendon progenitors during healing. We show that Axin2+ tenocytes readily expand in vitro and express stem cell markers. In vivo, Axin2+ cells are major functional contributors to repair: Axin2+ tenocytes de-differentiate, expand, and re-adopt a tenocyte fate post-injury. Specific loss of Wnt secretion in Axin2+ cells alters their stem cell identity and disrupts their activation upon injury, severely compromising healing. Our work highlights Axin2+ tenocytes as quiescent stem cells embedded in dense matrix, which are uniquely regulated in an autocrine manner and are central organizers of robust tendon healing.
2024-10-22 | GSE188598 | GEO