Restricting the ligation step of non-homologous end-joining

Non-homologous end-joining is an important pathway for repairing DNA double-strand breaks. The budding yeast Saccharomyces cerevisiae possesses two proteins, Nej1/Lif2 and Ntr1/Spp382, which play a role in restricting the activity of Dnl4-Lif1, the complex that executes the final ligation step of this process.     

S-Adenosyl-L-methionine-dependent restriction enzymes

Restriction-modification (R-M) enzymes are classified into type I, II, III, and IV, based on their recognition sequence, subunit composition, cleavage position, and cofactor requirements. While the role of S-Adenosyl-L-methionine (AdoMet) as the methyl group donor in the methylation reaction is undisputed, its requirement in DNA cleavage reaction has been subject to intense study. AdoMet is a prerequisite for the DNA cleavage by most type I enzymes known so far, with the exception of R.EcoR124I. A number of new type II restriction enzymes belonging to the type IIB and IIG family were found to show AdoMet dependence for their cleavage reaction. The type III enzymes have been found to require AdoMet for their restriction function. AdoMet functions as an allosteric effector of the DNA cleavage reaction and has been shown to bring about conformational changes in the protein upon binding.     

Retrovirus restriction factors

A number of cellular genes have recently been identified that actively inhibit retrovirus replication and so protect cells from infection. The genes target many distinct steps in the viral life cycle: entry, viral DNA synthesis, intracellular movement of viral nucleic acids, and viral gene expression. These restriction systems constitute newly appreciated components of an innate immunity that may be important for survival of a host exposed to retrovirus infection. It may someday be possible to enhance or activate these systems to induce antiviral states.     

Restricting HIV the SAMHD1 way: through nucleotide starvation

HIV replication is limited by cellular restriction factors, such as APOBEC and tetherin, which themselves are counteracted by viral proteins. SAMHD1 was recently identified as a novel HIV restriction factor in myeloid cells, and was shown to be blocked by the lentiviral protein Vpx. SAMHD1 limits viral replication through an original mechanism: it hydrolyses intracellular dNTPs in non-cycling cells, thus decreasing the amount of these key substrates, which are required for viral DNA synthesis. In this Progress article, we describe how SAMHD1 regulates the pool of intracellular nucleotides to control HIV replication and the innate immune response.     

Restricting environmental stimulation influences levels and variability of plasma cortisol

Restricting stimulation from the environment has been shown to alter psychological and physiological states. The present study of 27 healthy subjects examines the effects of restricted environmental stimulation technique (REST) on plasma levels of cortisol and variability in plasma cortisol levels across repeated REST sessions. The REST environment consisted of a 1.2 X 1.2 X 2.4-m ovoid chamber containing 25 cm of saturated MgSO4 solution (sp gr 1.28) maintained at 34.5 degrees F. The buoyant supinely floating subject experienced a minimum of light, sound, and temperature awareness and spatial orientation. The non-REST environment was a cushioned reclining chair in a quiet dimly lit room. The 5-wk protocol consisted of four visits for blood sampling during a 2-wk baseline followed by eight REST or non-REST sessions, 40 min each, with blood samples taken on four nonsession days between sessions 5 and 8. Variability in plasma cortisol was expressed in terms of standard deviation. REST was associated with across-session decreases of 21.6% in plasma cortisol and 50.5% in plasma cortisol variability, whereas no changes in these measures occurred in non-REST. It is concluded that REST influences both static and dynamic aspects of adrenocortical function, possibly altering the feedback monitoring of plasma cortisol.     

Construction of restriction maps

A computer program is described, which constructs maps of restrictionendonuclease cleavage sites in linear or circular DNA molecules,given the fragment lengths in single and double digestions withtwo enzymes. The algorithm is based upon a partition methodand a very simple rule to chain fragments. The program is writtenin Prolog II. Received on July 28, 1987; accepted on December 31, 1987     

Betaine improved restriction digestion

Here we report that supplementation of a common compound betaine (1-carboxy-N,N,N-trimethylmethanaminium inner salt) enhances restriction digestion of DNA molecules being resistant to digestion despite the existence of recognition sites. A previous study reported total isostabilization of DNA was achieved in the presence of 5.2M of betaine, however, we have observed the enhancement of restriction kinetics at 0.3M of betaine, therefore, it likely provided some catalytic proficiency to restriction enzymes rather than the induction of DNA conformational changes. Betaine also enhances catalytic efficiency of PCR, and our result of restriction digestion, taken together, suggests potential application of betaine in other enzymatic reactions in an aqueous solution.     

Protein restriction and cancer

Protein restriction without malnutrition is currently an effective nutritional intervention known to prevent diseases and promote health span from yeast to human. Recently, low protein diets are reported to be associated with lowered cancer incidence and mortality risk of cancers in human. In murine models, protein restriction inhibits tumor growth via mTOR signaling pathway. IGF-1, amino acid metabolic programing, FGF21, and autophagy may also serve as potential mechanisms of protein restriction mediated cancer prevention. Together, dietary intervention aimed at reducing protein intake can be beneficial and has the potential to be widely adopted and effective in preventing and treating cancers.     

Nucleoside triphosphate-dependent restriction enzymes

The known nucleoside triphosphate-dependent restriction enzymes are hetero-oligomeric proteins that behave as molecular machines in response to their target sequences. They translocate DNA in a process dependent on the hydrolysis of a nucleoside triphosphate. For the ATP-dependent type I and type III restriction and modification systems, the collision of translocating complexes triggers hydrolysis of phosphodiester bonds in unmodified DNA to generate double-strand breaks. Type I endonucleases break the DNA at unspecified sequences remote from the target sequence, type III endonucleases at a fixed position close to the target sequence. Type I and type III restriction and modification (R-M) systems are notable for effective post-translational control of their endonuclease activity. For some type I enzymes, this control is mediated by proteolytic degradation of that subunit of the complex which is essential for DNA translocation and breakage. This control, lacking in the well-studied type II R-M systems, provides extraordinarily effective protection of resident DNA should it acquire unmodified target sequences. The only well-documented GTP-dependent restriction enzyme, McrBC, requires methylated target sequences for the initiation of phosphodiester bond cleavage.     

Energy restriction with protein restriction increases basal metabolism and meal-induced thermogenesis in rats

We previously observed an increased sympathetic nervous system (SNS) activity that was partly responsible for a defect in the insulin secretion response to glucose after postweaning protein-energy restriction (PER) in female rats. These results, together with other data on low-protein feeding, suggested that a low protein-to-energy ratio (P/E) in the diet could stimulate energy expenditure (EE), but direct measurements of EE have never been reported under conditions of PER. The goal of the present study was thus to quantify the changes induced by PER to body composition, the various parameters of EE, and plasma triiodothyronine levels. PER induced severe growth retardation, but the subcutaneous white and interscapular brown adipose tissue masses were preserved. Basal metabolism, meal-induced thermogenesis, and triiodothyronine levels were increased, but substrate utilization by the working muscles was unaffected. Meal-induced thermogenesis was increased by spontaneous activity in PER rats only. These results suggest that rats adapt to a low P/E in the diet by burning part of their excess nonprotein energy and storing the remaining excess in subcutaneous adipose tissue.     

Inhibition of cytolytic lymphocytes by homologous restriction factor. Lack of species restriction

Homologous restriction factor (HRF) has been shown to inhibit complement-mediated lysis in a species-restrictive manner. Human HRF is able to block lysis by human complement but not by complement from other species. HRF has also been found in the membrane of lymphokine-activated killer (LAK) cells. When this HRF is inserted into sheep erythrocyte membranes, it is able to protect the erythrocyte from LAK cell lysis. In this report, we show that while HRF can inhibit human complement but not rat complement-mediated hemolysis, it is able to inhibit LAK cell lysis by both human and rat LAK cells. HRF is therefore a more general protective protein than has been previously thought.     

Restricting Apoptosis for Postmitotic Cell Survival and its Relevance to Cancer

The importance of apoptosis in multicellular organisms is signified by the high degree of genetic conservation in the core components of this pathway from C. elegans through mammals. However, as the cells which comprise these organisms have diversified and become more specialized, so have the mechanisms which regulate the apoptotic pathway. The complex regulatory mechanisms by which the apoptotic pathway is refined are perhaps most apparent in differentiated postmitotic cells such as neurons, cardiomyocytes, and skeletal myotubes. The lack of significant regenerative potential in postmitotic cells demands that they must persist long-term, often for the full lifespan of an organism. Recent studies have identified several diverse mechanisms by which postmitotic cells restrict their apoptotic potential. Importantly, these mechanisms may also be co-opted by cancer cells in order to evade apoptosis.     

Restricting hyperhydricity of lettuce using screen raft and water absorbent resin

A method for reducing hyperhydricity of lettuce shoots regenerated from culture on liquid and gelled media is described. In the case of liquid medium, hyperhydricity could be decreased by 30% by using a filter paper raft or a polyester screen raft to prevent immersion of inoculum in the culture medium. In the case of gelled medium, replacing agar with the water-absorbent resin SuperSorb could successfully prevent hyperhydricity without depressing organogenesis.