Sumoylation of Cas9 at lysine 848 regulates protein stability and DNA binding

CRISPR/Cas9 is a popular genome editing technology. Although widely used, little is known about how this prokaryotic system behaves in humans. An unwanted consequence of eukaryotic Cas9 expression is off-target DNA binding leading to mutagenesis. Safer clinical implementation of CRISPR/Cas9 necessitates a finer understanding of the regulatory mechanisms governing Cas9 behavior in humans. Here, we report our discovery of Cas9 sumoylation and ubiquitylation, the first post-translational modifications to be described on this enzyme. We found that the major SUMO2/3 conjugation site on Cas9 is K848, a key positively charged residue in the HNH nuclease domain that is known to interact with target DNA and contribute to off-target DNA binding. Our results suggest that Cas9 ubiquitylation leads to decreased stability via proteasomal degradation. Preventing Cas9 sumoylation through conversion of K848 into arginine or pharmacologic inhibition of cellular sumoylation enhances the enzyme’s turnover and diminishes guide RNA-directed DNA binding efficacy, suggesting that sumoylation at this site regulates Cas9 stability and DNA binding. More research is needed to fully understand the implications of these modifications for Cas9 specificity.

Cell wall deficiency as an escape mechanism from phage infection

The cell wall plays a central role in protecting bacteria from some environmental stresses, but not against all. In fact, in some cases, an elaborate cell envelope may even render the cell more vulnerable. For example, it contains molecules or complexes that bacteriophages recognize as the first step of host invasion, such as proteins and sugars, or cell appendages such as pili or flagella. In order to counteract phages, bacteria have evolved multiple escape mechanisms, such as restriction-modification, abortive infection, CRISPR/Cas systems or phage inhibitors. In this perspective review, we present the hypothesis that bacteria may have additional means to escape phage attack. Some bacteria are known to be able to shed their cell wall in response to environmental stresses, yielding cells that transiently lack a cell wall. In this wall-less state, the bacteria may be temporarily protected against phages, since they lack the essential entities that are necessary for phage binding and infection. Given that cell wall deficiency can be triggered by clinically administered antibiotics, phage escape could be an unwanted consequence that limits the use of phage therapy for treating stubborn infections.

Outsourcing problems or regulating altruism? Parliamentary debates on domestic and cross-border surrogacy in Finland and Norway

This article employs the concept of respectability and the discursive representation of gender equality policies to discuss how surrogacy is represented in Nordic parliamentary debates and policy documents. The article’s objective is to study how respectability, problems and equality are represented and discursively and rhetorically produced through a comparative study of Finnish and Norwegian political discourses on domestic unpaid surrogacy and cross-border commercial surrogacy. The article uses rhetorical and discursive analysis to analyse the Finnish and Norwegian Parliaments’ bills, members’ initiatives and proceedings from 2002 to 2018. Finland’s policy on surrogacy has evolved from an unregulated and permissive approach towards a more restrictive one, with discourses focusing on medicalisation, equality, altruism and safety concerning domestic surrogacy and problems and risks concerning cross-border surrogacy. Norway’s policy on surrogacy has been restrictive consistently, with discourses focusing on surrogacy as a transnational social problem involving exploitation of women and children, and biocentrism. Analysing surrogacy regulation in Nordic welfare states, the author concludes that policies and parliamentary debates in both countries have expressed expectations for inclusive health policies and social security for families. Cross-border surrogacy is characterised as an unwanted consequence of globalisation and marketisation of reproduction. Surrogate mothers’ respectability is constructed through rhetoric on differences in terms of nationality, class and binary representations of female caring and instrumentalism.

Erratum to: Generalizing the relativistic quantization condition to include all three-pion isospin channels

We have found an error in a statement following eq. (2.5) of our paper, concerning the function f (a, b, k) that first appears in that equation. The issue arises in the statement that it is convenient to take the function f (a, b, k) to be exchange symmetric with respect to its three arguments. This has the unwanted consequence of making six of the seven operators in the column vector of eq. (2.4) identically equal. This, in turn, implies that many operators are identically zero in the definite isospin basis, considered in section 2.4. To repair this, the last sentence of the paragraph containing eq. (2.4), starting “It is convenient for the subsequent…”, should be removed, as should footnote 3 and the next paragraph, beginning with “At this point, the reader may wonder why…”.

Consequences of assisted reproductive techniques on the embryonic epigenome in cattle

Procedures used in assisted reproduction have been under constant scrutiny since their inception with the goal of improving the number and quality of embryos produced. However, invitro production of embryos is not without complications because many fertilised oocytes fail to become blastocysts, and even those that do often differ in the genetic output compared with their invivo counterparts. Thus only a portion of those transferred complete normal fetal development. An unwanted consequence of bovine assisted reproductive technology (ART) is the induction of a syndrome characterised by fetal overgrowth and placental abnormalities, namely large offspring syndrome; a condition associated with inappropriate control of the epigenome. Epigenetics is the study of chromatin and its effects on genetic output. Establishment and maintenance of epigenetic marks during gametogenesis and embryogenesis is imperative for the maintenance of cell identity and function. ARTs are implemented during times of vast epigenetic reprogramming; as a result, many studies have identified ART-induced deviations in epigenetic regulation in mammalian gametes and embryos. This review describes the various layers of epigenetic regulation and discusses findings pertaining to the effects of ART on the epigenome of bovine gametes and the preimplantation embryo.

Status of Waqf Properties As Debt’s Collateral

The lack of understanding on land waqf creates many issues related to waqf itself, i.e. offers endowed land as deb’ts collateral. In Article 40 alphabeth (a) Law Number 41 Year 2004 on Waqf, there is prohibition to include waqf properties as collateral. Waqf properties’ status change that inchorent with Law Number 41 Year 2004 on Waqf, one of it is including land waqf as debt’s collateral will cause unwanted consequence on the collateral agreement itself. In fact, there are still violations on Article 40 alphabeth (a) Law Number 41 Year 2004 on Waqf. These situations are caused by minimum awareness on management of waqf properties from nadzir, heirs and bank as the creditor towards implementation of waqf properties’ status change. Therefore, consequence of waqf properties’ status change that violates Article 40 alphabeth (a) Law Number 41 Year 2004 on Waqf will not immediately null and void; however, debt’s repayment still needs to be fulfilled by debtor corresponds with Article 1131 BW. It required efforts to minimize issues on waqf through socialization of waqf towards nadzir, heirs, bank as creditor and wide community, especially waqf properties’ status change so there will not be any issues on implementation and management of waqf properties in the future.

The problem of nitrogen disposal in the obese

Amino-N is preserved because of the scarcity and nutritional importance of protein. Excretion requires its conversion to ammonia, later incorporated into urea. Under conditions of excess dietary energy, the body cannot easily dispose of the excess amino-N against the evolutively adapted schemes that prevent its wastage; thus ammonia and glutamine formation (and urea excretion) are decreased. High lipid (and energy) availability limits the utilisation of glucose, and high glucose spares the production of ammonium from amino acids, limiting the synthesis of glutamine and its utilisation by the intestine and kidney. The amino acid composition of the diet affects the production of ammonium depending on its composition and the individual amino acid catabolic pathways. Surplus amino acids enhance protein synthesis and growth, and the synthesis of non-protein-N-containing compounds. But these outlets are not enough; consequently, less-conventional mechanisms are activated, such as increased synthesis of NO∙ followed by higher nitrite (and nitrate) excretion and changes in the microbiota. There is also a significant production of N2 gas, through unknown mechanisms. Health consequences of amino-N surplus are difficult to fathom because of the sparse data available, but it can be speculated that the effects may be negative, largely because the fundamental N homeostasis is stretched out of normalcy, forcing the N removal through pathways unprepared for that task. The unreliable results of hyperproteic diets, and part of the dysregulation found in the metabolic syndrome may be an unwanted consequence of this N disposal conflict.

The Stem Cell Hypothesis of Aging

BACKGROUND: There is probably no single way to age. Indeed, so far there is no single accepted explanation or mechanisms of aging (although more than 300 theories have been proposed). There is an overall decline in tissue regenerative potential with age, and the question arises as to whether this is due to the intrinsic aging of stem cells or rather to the impairment of stem cell function in the aged tissue environment.CONTENT: Recent data suggest that we age, in part, because our self-renewing stem cells grow old as a result of heritable intrinsic events, such as DNA damage, as well as extrinsic forces, such as changes in their supporting niches. Mechanisms that suppress the development of cancer, such as senescence and apoptosis, which rely on telomere shortening and the activities of p53 and p16INK4a may also induce an unwanted consequence: a decline in the replicative function of certain stem cells types with advancing age. This decrease regenerative capacity appears to pointing to the stem cell hypothesis of aging.SUMMARY: Recent evidence suggested that we grow old partly because of our stem cells grow old as a result of mechanisms that suppress the development of cancer over a lifetime. We believe that a further, more precise mechanistic understanding of this process will be required before this knowledge can be translated into human anti-aging therapies.KEYWORDS: stem cells, senescence, telomere, DNA damage, epigenetic, aging

Global antibacterial resistance issues

The clinical use of antimicrobial agents has spawned, as an unwanted consequence, the widespread emergence of bacteria resistant to these valuable drugs. During the past fifty years, resistant organisms have caused problems throughout the world, as will be documented in this brief paper.