RESUMO
Translin and translin-associated factor-x are highly conserved in eukaroytes; they can form heteromeric complexes (known as C3POs) and participate in various nucleic acid metabolism pathways. In humans and Drosophila, C3POs cleave the fragmented siRNA passenger strands and facilitate the activation of RNA-induced silencing complex, the effector complex of RNA interference (RNAi). Here, we report three crystal structures of Nanoarchaeum equitans (Ne) C3PO. The apo-NeC3PO structure adopts an open form and unravels a potential substrates entryway for the first time. The NeC3PO:ssRNA and NeC3PO:ssDNA complexes fold like closed football with the substrates captured at the inner cavities. The NeC3PO:ssRNA structure represents the only catalytic form C3PO complex available to date; with mutagenesis and in vitro cleavage assays, the structure provides critical insights into the substrate binding and the two-cation-assisted catalytic mechanisms that are shared by eukaryotic C3POs. The work presented here further advances our understanding on the RNAi pathway.
Assuntos
Proteínas Arqueais/química , Proteínas Arqueais/metabolismo , Modelos Moleculares , Conformação Molecular , Clivagem do RNA , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , RNA/química , Sítios de Ligação , Cátions/química , Ligação Proteica , RNA/metabolismo , Ribonucleases/química , Ribonucleases/metabolismo , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
Kabuki syndrome (KS) is a rare genetic disorder typically characterized by facial abnormalities, developmental delay, cognitive dysfunction, and organ impairment. In this report, fibroblast cells obtained from a KS patient containing a heterozygous KMT2D c.12592 C>T mutation (p.R4198X) were reprogrammed using non-integrative Sendai virus to generate three induced pluripotent stem cell (iPSC) clones. The iPSC lines retained the KS patient mutation, and displayed normal karyotypes, pluripotency marker expression, and the ability to differentiate into the three germ layers.
Assuntos
Doenças Hematológicas , Células-Tronco Pluripotentes Induzidas , Doenças Vestibulares , Anormalidades Múltiplas , Face/anormalidades , Doenças Hematológicas/genética , Humanos , Mutação/genética , Doenças Vestibulares/genéticaRESUMO
For cancer cells to survive during extracellular matrix (ECM) detachment, they must inhibit anoikis and rectify metabolic deficiencies that cause non-apoptotic cell death. Previous studies in ECM-detached cells have linked non-apoptotic cell death to reactive oxygen species (ROS) generation, although the mechanistic underpinnings of this link remain poorly defined. Here, we uncover a role for receptor-interacting protein kinase 1 (RIPK1) in the modulation of ROS and cell viability during ECM detachment. We find that RIPK1 activation during ECM detachment results in mitophagy induction through a mechanism dependent on the mitochondrial phosphatase PGAM5. As a consequence of mitophagy, ECM-detached cells experience diminished NADPH production in the mitochondria, and the subsequent elevation in ROS levels leads to non-apoptotic death. Furthermore, we find that antagonizing RIPK1/PGAM5 enhances tumour formation in vivo. Thus, RIPK1-mediated induction of mitophagy may be an efficacious target for therapeutics aimed at eliminating ECM-detached cancer cells.