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In vitro–transcribed guide RNAs trigger an innate immune response via the RIG-I pathway

PLOS Biology (new articles) - Mon, 2018-07-16 23:00

by Beeke Wienert, Jiyung Shin, Elena Zelin, Kathleen Pestal, Jacob E. Corn

Clustered, regularly interspaced, short palindromic repeat (CRISPR)–CRISPR-associated 9 (Cas9) genome editing is revolutionizing fundamental research and has great potential for the treatment of many diseases. While editing of immortalized cell lines has become relatively easy, editing of therapeutically relevant primary cells and tissues can remain challenging. One recent advancement is the delivery of a Cas9 protein and an in vitro–transcribed (IVT) guide RNA (gRNA) as a precomplexed ribonucleoprotein (RNP). This approach allows editing of primary cells such as T cells and hematopoietic stem cells, but the consequences beyond genome editing of introducing foreign Cas9 RNPs into mammalian cells are not fully understood. Here, we show that the IVT gRNAs commonly used by many laboratories for RNP editing trigger a potent innate immune response that is similar to canonical immune-stimulating ligands. IVT gRNAs are recognized in the cytosol through the retinoic acid–inducible gene I (RIG-I) pathway but not the melanoma differentiation–associated gene 5 (MDA5) pathway, thereby triggering a type I interferon response. Removal of the 5’-triphosphate from gRNAs ameliorates inflammatory signaling and prevents the loss of viability associated with genome editing in hematopoietic stem cells. The potential for Cas9 RNP editing to induce a potent antiviral response indicates that care must be taken when designing therapeutic strategies to edit primary cells.
Categories: Biology, Journals

Inter-subunit interactions drive divergent dynamics in mammalian and <i>Plasmodium</i> actin filaments

PLOS Biology (new articles) - Mon, 2018-07-16 23:00

by Ross G. Douglas, Prajwal Nandekar, Julia-Elisabeth Aktories, Hirdesh Kumar, Rebekka Weber, Julia M. Sattler, Mirko Singer, Simone Lepper, S. Kashif Sadiq, Rebecca C. Wade, Friedrich Frischknecht

Cell motility is essential for protozoan and metazoan organisms and typically relies on the dynamic turnover of actin filaments. In metazoans, monomeric actin polymerises into usually long and stable filaments, while some protozoans form only short and highly dynamic actin filaments. These different dynamics are partly due to the different sets of actin regulatory proteins and partly due to the sequence of actin itself. Here we probe the interactions of actin subunits within divergent actin filaments using a comparative dynamic molecular model and explore their functions using Plasmodium, the protozoan causing malaria, and mouse melanoma derived B16-F1 cells as model systems. Parasite actin tagged to a fluorescent protein (FP) did not incorporate into mammalian actin filaments, and rabbit actin-FP did not incorporate into parasite actin filaments. However, exchanging the most divergent region of actin subdomain 3 allowed such reciprocal incorporation. The exchange of a single amino acid residue in subdomain 2 (N41H) of Plasmodium actin markedly improved incorporation into mammalian filaments. In the parasite, modification of most subunit–subunit interaction sites was lethal, whereas changes in actin subdomains 1 and 4 reduced efficient parasite motility and hence mosquito organ penetration. The strong penetration defects could be rescued by overexpression of the actin filament regulator coronin. Through these comparative approaches we identified an essential and common contributor, subdomain 3, which drives the differential dynamic behaviour of two highly divergent eukaryotic actins in motile cells.
Categories: Biology, Journals

Stone Age bakers made first bread thousands of years before farming

Genetics - Mon, 2018-07-16 21:00
Evidence of the first early bread suggests humans were baking with wheat and oats thousands of years before they began farming the cereals
Categories: Biology

Stone Age bakers made first bread thousands of years before farming

HIV and AIDS - Mon, 2018-07-16 21:00
Evidence of the first early bread suggests humans were baking with wheat and oats thousands of years before they began farming the cereals

Stone Age bakers made first bread thousands of years before farming

Genetically modified organisms (GMO) - Mon, 2018-07-16 21:00
Evidence of the first early bread suggests humans were baking with wheat and oats thousands of years before they began farming the cereals
Categories: Science and society

UK announces plans for two spaceports that could see launches by 2021

Genetics - Mon, 2018-07-16 18:51
The UK is funding the development of spaceports in Scotland and Cornwall, but plans for US firms to launch small satellites could run afoul of US export laws
Categories: Biology

UK announces plans for two spaceports that could see launches by 2021

HIV and AIDS - Mon, 2018-07-16 18:51
The UK is funding the development of spaceports in Scotland and Cornwall, but plans for US firms to launch small satellites could run afoul of US export laws

UK announces plans for two spaceports that could see launches by 2021

Genetically modified organisms (GMO) - Mon, 2018-07-16 18:51
The UK is funding the development of spaceports in Scotland and Cornwall, but plans for US firms to launch small satellites could run afoul of US export laws
Categories: Science and society

Wireless implant lights up inside the body to kill cancer

Genetics - Mon, 2018-07-16 18:00
Researchers have developed a sticky sheet that could allow a wirelessly-powered LED chip to be stuck inside the body to deliver "photodynamic therapy"
Categories: Biology

CRISPR gene editing is not quite as precise and as safe as thought

Genetics - Mon, 2018-07-16 18:00
A study has found that CRISPR can delete large chunks of DNA, suggesting it could cause cancer if used to treat diseases by editing many cells in the body
Categories: Biology

Wireless implant lights up inside the body to kill cancer

HIV and AIDS - Mon, 2018-07-16 18:00
Researchers have developed a sticky sheet that could allow a wirelessly-powered LED chip to be stuck inside the body to deliver "photodynamic therapy"

CRISPR gene editing is not quite as precise and as safe as thought

HIV and AIDS - Mon, 2018-07-16 18:00
A study has found that CRISPR can delete large chunks of DNA, suggesting it could cause cancer if used to treat diseases by editing many cells in the body

Wireless implant lights up inside the body to kill cancer

Genetically modified organisms (GMO) - Mon, 2018-07-16 18:00
Researchers have developed a sticky sheet that could allow a wirelessly-powered LED chip to be stuck inside the body to deliver "photodynamic therapy"
Categories: Science and society

CRISPR gene editing is not quite as precise and as safe as thought

Genetically modified organisms (GMO) - Mon, 2018-07-16 18:00
A study has found that CRISPR can delete large chunks of DNA, suggesting it could cause cancer if used to treat diseases by editing many cells in the body
Categories: Science and society

Thought plastic was bad enough? Here’s another reason to worry

Genetically modified organisms (GMO) - Mon, 2018-07-16 14:00
We knew marine plastic strangles birds and poisons fish. But it can also pick up some deadly hitch-hikers – with even more profound consequences
Categories: Science and society

Thought plastic was bad enough? Here’s another reason to worry

Genetics - Mon, 2018-07-16 14:00
We knew marine plastic strangles birds and poisons fish. But it can also pick up some deadly hitch-hikers – with even more profound consequences
Categories: Biology

A no-deal Brexit could lead to a catastrophe for science in the UK

Genetics - Mon, 2018-07-16 12:00
The flurry of cabinet resignations in the aftermath of the Chequers agreement leaves the UK at serious risk of crashing out of the EU without a deal
Categories: Biology

Honeybees gang up to roast invading hornets alive — at a terrible cost

Genetics - Mon, 2018-07-16 12:00
The worker bees that form “hot defensive bee balls” are effectively kamikaze fighters, with the heat from the ball shortening their life expectancy
Categories: Biology

A no-deal Brexit could lead to a catastrophe for science in the UK

Genetically modified organisms (GMO) - Mon, 2018-07-16 12:00
The flurry of cabinet resignations in the aftermath of the Chequers agreement leaves the UK at serious risk of crashing out of the EU without a deal
Categories: Science and society

Honeybees gang up to roast invading hornets alive — at a terrible cost

Genetically modified organisms (GMO) - Mon, 2018-07-16 12:00
The worker bees that form “hot defensive bee balls” are effectively kamikaze fighters, with the heat from the ball shortening their life expectancy
Categories: Science and society