Agriculture, Forestry, and Other Land Use (AFOLU) is unique among the sectors considered in this volume, since the mitigation potential is derived from both an enhancement of removals of greenhouse gases (GHG), as well as reduction of emissions through management of land and livestock (robust evidence; high agreement). The land provides food that feeds the Earth’s human population of ca. 7 billion, fibre for a variety of purposes, livelihoods for billions of people worldwide, and is a critical resource for sustainable development in many regions. Agriculture is frequently central to the livelihoods of many social groups, especially in developing countries where it often accounts for a significant share of production. In addition to food and fibre, the land provides a multitude of ecosystem services; climate change mitigation is just one of many that are vital to human well-being (robust evidence; high agreement). Mitigation options in the AFOLU sector, therefore, need to be assessed, as far as possible, for their potential impact on all other services provided by land. [Section 11.1]

/pdf/agriculture-forestry-and-other-land-use-afolu-4vwhm7hk8n.pdf

Agriculture, Forestry and Other Land Use (AFOLU)

The present disclosure provides a DNA-targeting RNA that comprises a targeting sequence and, together with a modifying polypeptide, provides for site-specific modification of a target DNA and/or a polypeptide associated with the target DNA. The present disclosure further provides site-specific modifying polypeptides. The present disclosure further provides methods of site-specific modification of a target DNA and/or a polypeptide associated with the target DNA The present disclosure provides methods of modulating transcription of a target nucleic acid in a target cell, generally involving contacting the target nucleic acid with an enzymatically inactive Cas9 polypeptide and a DNA-targeting RNA. Kits and compositions for carrying out the methods are also provided. The present disclosure provides genetically modified cells that produce Cas9; and Cas9 transgenic non-human multicellular organisms.

Methods and compositions for rna-directed target dna modification and for rna-directed modulation of transcription

Bioenergy deployment offers significant potential for climate change mitigation, but also carries considerable risks. In this review, we bring together perspectives of various communities involved in the research and regulation of bioenergy deployment in the context of climate change mitigation: Land-use and energy experts, land-use and integrated assessment modelers, human geographers, ecosystem researchers, climate scientists and two different strands of life-cycle assessment experts. We summarize technological options, outline the state-of-the-art knowledge on various climate effects, provide an update on estimates of technical resource potential and comprehensively identify sustainability effects. Cellulosic feedstocks, increased end-use efficiency, improved land carbon-stock management and residue use, and, when fully developed, BECCS appear as the most promising options, depending on development costs, implementation, learning, and risk management. Combined heat and power, efficient biomass cookstoves and small-scale power generation for rural areas can help to promote energy access and sustainable development, along with reduced emissions. We estimate the sustainable technical potential as up to 100EJ: high agreement; 100-300EJ: medium agreement; above 300EJ: low agreement. Stabilization scenarios indicate that bioenergy may supply from 10 to 245EJyr(-1) to global primary energy supply by 2050. Models indicate that, if technological and governance preconditions are met, large-scale deployment (>200EJ), together with BECCS, could help to keep global warming below 2 degrees degrees of preindustrial levels; but such high deployment of land-intensive bioenergy feedstocks could also lead to detrimental climate effects, negatively impact ecosystems, biodiversity and livelihoods. The integration of bioenergy systems into agriculture and forest landscapes can improve land and water use efficiency and help address concerns about environmental impacts. We conclude that the high variability in pathways, uncertainties in technological development and ambiguity in political decision render forecasts on deployment levels and climate effects very difficult. However, uncertainty about projections should not preclude pursuing beneficial bioenergy options.

/pdf/bioenergy-and-climate-change-mitigation-an-assessment-4kmc9b5mav.pdf

Bioenergy and climate change mitigation: an assessment.

The recombinant HIV-1 Tat protein contains a small region corresponding to residues 47YGRKKRRQRR57R, which is capable of translocating cargoes of different molecular sizes, such as proteins, DNA, RNA, or drugs, across the cell membrane in an apparently energy-independent manner. The pathway that these peptides follow for entry into the cell has been the subject of strong controversy for the last decade. This peptide is highly basic and hydrophilic. Therefore, a central question that any candidate mechanism has to answer is how this highly hydrophilic peptide is able to cross the hydrophobic barrier imposed by the cell membrane. We propose a mechanism for the spontaneous translocation of the Tat peptides across a lipid membrane. This mechanism involves strong interactions between the Tat peptides and the phosphate groups on both sides of the lipid bilayer, the insertion of charged side chains that nucleate the formation of a transient pore, followed by the translocation of the Tat peptides by diffusing on the pore surface. This mechanism explains how key ingredients, such as the cooperativity among the peptides, the large positive charge, and specifically the arginine amino acids, contribute to the uptake. The proposed mechanism also illustrates the importance of membrane fluctuations. Indeed, mechanisms that involve large fluctuations of the membrane structure, such as transient pores and the insertion of charged amino acid side chains, may be common and perhaps central to the functions of many membrane protein functions.

https://www.pnas.org/content/pnas/104/52/20805.full.pdf

Molecular dynamics simulations suggest a mechanism for translocation of the HIV-1 TAT peptide across lipid membranes

Antimicrobial peptides (AMPs): Ancient compounds that represent novel weapons in the fight against bacteria.

Novel classes and applications of cell-penetrating peptides (CPPs) are being constantly discovered since they were first identified 2 decades ago. These short cationic peptides (nanomolecules) either by covalent binding or by noncovalent binding can traverse cell membranes and deliver a variety of molecules that are unable to overcome the permeability barrier in their own capacity. The ability of the CPPs to deliver variety of macromolecules, such as oligonucleotides, therapeutic drugs, proteins, and medical imaging agents, by forming nanoparticulate carriers in a range of cells has led them to emerge as a potential tool for both macromolecule delivery application and to gain insight into the fundamentals of mechanism of cellular uptake across the plasma membrane. This review explores the recent advances, challenges, and future prospects in the field of CPP-mediated cargo delivery in mammalian and plant cells. Studies have been conducted into the peptide chemistry and stability of CPP-macromolecular complexes. Most of the CPPs have been shown to be nontoxic and do not interfere with the functionality of the macromolecules delivered across the cell membrane. The mechanism of uptake of CPP-cargo complexes and the uptake of CPPs alone across the plasma membrane remains unresolved. As the world of CPPs is rapidly advancing in both mammalian and plant system, there is a promising future for the various applications of transduction and transfection into intact cells.

https://iubmb.onlinelibrary.wiley.com/doi/pdf/10.1002/iub.297

Cell-penetrating peptides: Nanocarrier for macromolecule delivery in living cells

Somatic embryogenesis is a remarkable illustration of the dictum of plant totipotency During embryogenic induction of cells, there is differential gene expression resulting in synthesis of new mRNAs and proteins This genetic information in turn elicits diverse cellular and physiological responses that are involved in 'switching over' of the developmental programme of the somatic cells Various model systems have been widely investigated to understand the mechanisms of gene regulation during this developmental process, and an array of genes activated or differentially expressed during somatic embryogenesis have been isolated employing various molecular techniques Nonetheless, the precise mechanisms controlling plant gene expression and the detailed steps by which these genes direct the plant-specific process of somatic embryogenesis remain far from being clearly understood Thus, future trends involve characterization of development-specific genes during somatic embryogenesis to provide a deeper insight in understanding the mechanisms involved during differentiation of somatic cells and phenotypic expression of cellular totipotency in higher plants

Gene expression during somatic embryogenesis - recent advances

Microspore culture is contributing significantly in the field of plant breeding for crop improvement in general and cereals, in particular. In the present study, we investigated the uptake of fluorescently labeled cell-penetrating peptides (CPP; Tat, Tat2, M-Tat, peptide vascular endothelial-cadherin, transportan) in the freshly isolated triticale microspores (mid-late uninucleate stage). We demonstrated that Tat (RKKRRQRRR) and Tat2 (RKKRRQRRRRKKRRQRRR) are able to efficiently transduce GUS enzyme (272 kDa) in its functional form in 5 and 14% of the microspores, respectively, in a noncovalent manner. Pep-1, a synthetic CPP, was able to transduce GUS enzyme in its active form in 31% of the microspores. The effect of various endocytic and macropinocytic inhibitors on Tat2-mediated GUS enzyme delivery was studied and revealed a preferred micropinocytosis entry. DNase I protection assay and confocal laser microscopy was carried out to recommend a ratio of 4:1 Tat2-linear plasmid DNA (pActGUS) in complex preparation for microspore transfection. We further show that Tat2 can successfully deliver GUS gene in near to 2% triticale microspores. The negative control mutated Tat (M-Tat: AKKRRQRRR) failed to transducer the GUS protein and transfect the GUS gene in microspore nucleus. The ability of CPPs to deliver macromolecules (protein as well as linear plasmid DNA) noncovalently has been demonstrated in triticale isolated microspores. It further confirms potential applications of CPPs in developing simple, time saving, cost effective plant genetic engineering technologies.

Translocation of cell-penetrating peptides and delivery of their cargoes in triticale microspores.

The present invention provides a novel method for the transduction and/or transfection of plant cells. Cell-penetrating peptides (CPPs) have been successfully employed as nanocarriers to deliver proteins and oligonucleotides to single plant cell microspores as well as multi-cellular zygotic embryos. The efficiency of CPP internalization and further delivery of a macromolecular cargo comprising a protein and/or an oligonucleotide can be enhanced by permeabilization of the zygotic embryos.

Nanocarrier based plant transfection and transduction

Internalization of fluorescently labeled CPPs, pVEC, transportan and scrambled pVEC, in a range of plant cells was investigated. Cellular uptake of the peptides was found to be tissue dependent. pVEC and transportan were distinctly internalized in triticale mesophyll protoplasts, onion epidermal cells, leaf bases and root tips of seven-day old triticale seedlings but showed negligible florescence in coleoptile and leaf tips as observed under a fluorescence microscope. Further, pVEC and transportan uptake studies were focused on mesophyll protoplasts as a system of investigation. In fluorimetric studies transportan showed 2.3 times higher cellular internalization than pVEC in protoplasts, whereas scrambled pVEC failed to show any significant fluorescence. Effect of various factors on cellular internalization of pVEC and transportan in protoplasts was also investigated. The cellular uptake of both the peptides was concentration dependent and nonsaturable. The cellular uptake of pVEC and transportan was enhanced at low temperature (4 degrees C). The presence of endocytic/macropinocytosis inhibitors did not reduce the cellular uptake of the peptides, suggesting direct cell penetration, receptor-independent internalization of pVEC and transportan into the plant cells.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/psc.937

Archana Chugh

Papers

Cell-penetrating peptides: Nanocarrier for macromolecule delivery in living cells

Gene expression during somatic embryogenesis - recent advances

Translocation of cell-penetrating peptides and delivery of their cargoes in triticale microspores.

Nanocarrier based plant transfection and transduction

Cellular uptake of cell-penetrating peptides pVEC and transportan in plants.