CRISPR/Cas9 small promoter deletion in H19 lncRNA is associated with altered cell morphology and proliferation
Autor: | Mayara Magna de Lima Melo, Cristiana Libardi Miranda Furtado, Cláudia Pessoa, Daniel Pascoalino Pinheiro, Renan da Silva Santos, Ronald Feitosa Pinheiro, Gilvan Pessoa Furtado, Maria Claudia dos Santos Luciano, Sarah Leyenne Alves Sales, Kaio César Simiano Tavares, Louhanna Pinheiro Rodrigues Teixeira |
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Rok vydání: | 2021 |
Předmět: |
Carcinogenesis
Science Biology Cell morphology medicine.disease_cause Article Mice Cancer epigenetics Transcription (biology) Neoplasms Gene expression medicine Biomarkers Tumor Animals Humans Promoter Regions Genetic Gene Cell Proliferation Sequence Deletion Gene Editing Gene knockdown Multidisciplinary Base Sequence Cell Cycle Promoter female genital diseases and pregnancy complications Cell biology Mechanisms of disease Gene Knockdown Techniques Cancer cell embryonic structures Cytogenetic Analysis Medicine RNA Long Noncoding CRISPR-Cas Systems |
Zdroj: | Scientific Reports Scientific Reports, Vol 11, Iss 1, Pp 1-10 (2021) |
ISSN: | 2045-2322 |
Popis: | The imprinted H19 long non-coding RNA, a knowing oncofetal gene, presents a controversial role during the carcinogenesis process since its tumor suppressor or oncogenic activity is not completely elucidated. Since H19 lncRNA is involved in many biological pathways related to tumorigenesis, we sought to develop a non-cancer lineage with CRISPR-Cas9-mediated H19 knockdown (H19-) and observe the changes in a cellular context. To edit the promoter region of H19, two RNA guides were designed, and the murine C2C12 myoblast cells were transfected. H19 deletion was determined by DNA sequencing and gene expression by qPCR. We observed a small deletion (~ 60 bp) in the promoter region that presented four predicted transcription binding sites. The deletion reduced H19 expression (30%) and resulted in increased proliferative activity, altered morphological patterns including cell size and intracellular granularity, without changes in viability. The increased proliferation rate in the H19- cell seems to facilitate chromosomal abnormalities. The H19- myoblast presented characteristics similar to cancer cells, therefore the H19 lncRNA may be an important gene during the initiation of the tumorigenic process. Due to CRISPR/Cas9 permanent edition, the C2C12 H19- knockdown cells allows functional studies of H19 roles in tumorigenesis, prognosis, metastases, as well as drug resistance and targeted therapy. |
Databáze: | OpenAIRE |
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