Unraveling the 'DEAD-box' helicases of Plasmodium falciparum

The causative agent for the most fatal form of malaria, Plasmodium falciparum, has developed insecticide and drug resistance with time. Therefore combating this disease is becoming increasingly difficult and this calls for finding alternate ways to control malaria. One of the feasible ways could be to find out inhibitors/drugs specific for the indispensable enzymes of malaria parasite such as helicases. These helicases, which contain intrinsic nucleic acid-dependent ATPase activity, are capable of enzymatically unwinding energetically stable duplex nucleic acids into single-stranded templates and are required for all the nucleic acid transactions. Most of the helicases contain a set of nine extremely conserved amino acid sequences, which are called 'helicase motifs'. Due to the presence of the DEAD (Asp-Glu-Ala-Asp) in one of the conserved motifs, this family is also known as the 'DEAD-box' family. In this review, using bioinformatic approach, we describe the 'DEAD-box' helicases of malaria parasite P. falciparum. An in depth analysis shows that the parasite contains 22 full-length genes, some of which are homologues of well-characterized helicases of this family from other organisms. Recently we have cloned and characterized the first member of this family, which is a homologue of p68 and is expressed during the schizont stage of the development of the parasite [Pradhan, A., Chauhan, V.S., Tuteja, R., 2005a. A novel 'DEAD-box' DNA helicase from Plasmodium falciparum is homologous to p68. Mol. Biochem. Parasitol. 140, 55-60.; Pradhan A., Chauhan V.S., Tuteja R., 2005b. Plasmodium falciparum DNA helicase 60 is a schizont stage specific, bipolar and dual helicase stimulated by PKC phosphorylation. Mol. Biochem. Parasitol. 144, 133-141.]. It will be really interesting to clone and characterize other members of the 'DEAD-box' family and understand their role in the replication and transmission of the parasite. These detailed studies may help to identify a parasite-specific enzyme, which could be a potential drug target to treat malaria. The various steps at which this probable drug can act are also discussed.     

Unraveling DNA helicases from plant cells*

Plant Molecular Biology -     

Unraveling DNA helicases. Motif, structure, mechanism and function.

DNA helicases are molecular 'motor' enzymes that use the energy of NTP hydrolysis to separate transiently energetically stable duplex DNA into single strands. They are therefore essential in nearly all DNA metabolic transactions. They act as essential molecular tools for the cellular machinery. Since the discovery of the first DNA helicase in Escherichia coli in 1976, several have been isolated from both prokaryotic and eukaryotic systems. DNA helicases generally bind to ssDNA or ssDNA/dsDNA junctions and translocate mainly unidirectionally along the bound strand and disrupt the hydrogen bonds between the duplexes. Most helicases contain conserved motifs which act as an engine to drive DNA unwinding. Crystal structures have revealed an underlying common structural fold for their function. These structures suggest the role of the helicase motifs in catalytic function and offer clues as to how these proteins can translocate and unwind DNA. The genes containing helicase motifs may have evolved from a common ancestor. In this review we cover the conserved motifs, structural information, mechanism of DNA unwinding and translocation, and functional aspects of DNA helicases.     

The role of RecQ helicases in non-homologous end-joining

Abstract

DNA double-strand breaks are highly toxic DNA lesions that cause genomic instability, if not efficiently repaired. RecQ helicases are a family of highly conserved proteins that maintain genomic stability through their important roles in several DNA repair pathways, including DNA double-strand break repair. Double-strand breaks can be repaired by homologous recombination (HR) using sister chromatids as templates to facilitate precise DNA repair, or by an HR-independent mechanism known as non-homologous end-joining (NHEJ) (error-prone). NHEJ is a non-templated DNA repair process, in which DNA termini are directly ligated. Canonical NHEJ requires DNA-PKcs and Ku70/80, while alternative NHEJ pathways are DNA-PKcs and Ku70/80 independent. This review discusses the role of RecQ helicases in NHEJ, alternative (or back-up) NHEJ (B-NHEJ) and microhomology-mediated end-joining (MMEJ) in V(D)J recombination, class switch recombination and telomere maintenance.     

Unraveling the role of myosin in forming autophagosomes

Macroautophagy (hereafter autophagy) is a membrane-mediated catabolic process that occurs in response to a variety of intra- and extra-cellular stresses. It is characterized by the formation of specialized double-membrane vesicles, autophagosomes, which engulf organelles and long-lived proteins, and in turn fuse with lysosomes for degradation and recycling. How autophagosomes emerge is still unclear. The Atg1 kinase plays a crucial role in the induction of autophagosome formation. While several Atg (autophagy-related) proteins have been associated with, and have been found to regulate, Atg1 kinase activity, the downstream targets of Atg1 that trigger autophagy remain unknown. Our recent studies have identified a myosin light chain kinase (MLCK)-like kinase as the Atg1 kinase effector that induces the activation of myosin II, and have found it to be required for autophagosome formation during nutrient deprivation. We further demonstrated that Atg1-mediated myosin II activation is crucial for the movement of the Atg9 transmembrane protein between the Golgi and the forming autophagosome, which provides a membrane source for the formation of autophagosomes during starvation.     

Unraveling the role of myosin in forming autophagosomes

Macroautophagy (hereafter autophagy) is a membrane-mediated catabolic process that occurs in response to a variety of intra- and extra-cellular stresses. It is characterized by the formation of specialized double-membrane vesicles, autophagosomes, which engulf organelles and long-lived proteins, and in turn fuse with lysosomes for degradation and recycling. How autophagosomes emerge is still unclear. The Atg1 kinase plays a crucial role in the induction of autophagosome formation. While several Atg (autophagy-related) proteins have been associated with, and have been found to regulate, Atg1 kinase activity, the downstream targets of Atg1 that trigger autophagy remain unknown. Our recent studies have identified a myosin light chain kinase (MLCK)-like kinase as the Atg1 kinase effector that induces the activation of myosin II, and have found it to be required for autophagosome formation during nutrient deprivation. We further demonstrated that Atg1-mediated myosin II activation is crucial for the movement of the Atg9 transmembrane protein between the Golgi and the forming autophagosome, which provides a membrane source for the formation of autophagosomes during starvation.     

Unraveling the role of motoneuron autophagy in ALS

In recent years, the role of autophagy in the pathogenesis of most neurodegenerative diseases has transitioned into a limbo of protective or detrimental effects. Genetic evidence indicates that mutations in autophagy-regulatory genes can result in the occurrence of amyotrophic lateral sclerosis (ALS), suggesting a physiological role of the pathway to motoneuron function. However, experimental manipulation of autophagy in ALS models led to conflicting results depending on the intervention strategy and the disease model used. A recent work by the Maniatis group systematically explored the role of cell-specific autophagy in motoneurons at different disease stages, revealing surprising and unexpected findings. Autophagy activity at early stages may contribute to maintaining the structure and function of neuromuscular junctions, whereas at later steps of the disease it has a pathogenic activity possibly involving cell-nonautonomous mechanisms related to glial activation. This new study adds a new layer of complexity in the field, suggesting an intricate interplay between proteostasis alterations, the time-differential function of autophagy in neurons, and muscle innervation in ALS.     

Analysis of the role of RecQ helicases in RNAi in mammals

The identity of mammalian genes involved in RNA interference (RNAi), the targeted sequence-specific mRNA degradation by double-stranded RNA (dsRNA), is poorly defined. Here we report the analysis of mice with null mutations of Wrn, Blm, and RecQ1 genes that are related to Mut-7 and Qde3, two genes essential for RNAi in Caenorhabditis elegans and quelling in Neurospora, respectively. Our results suggest that Wrn, Blm, and RecQ1 are not involved in sequence-specific mRNA degradation in mammals in response to dsRNA, suggesting potential differences in the mammalian RNAi pathway.     

Unraveling the role of thiosulfate sulfurtransferase in metabolic diseases

Thiosulfate sulfurtransferase (TST, EC 2.8.1.1), also known as Rhodanese, is a mitochondrial enzyme which catalyzes the transfer of sulfur in several molecular pathways. After its initial identification as a cyanide detoxification enzyme, it was found that its functions also include sulfur metabolism, modification of iron‑sulfur clusters and the reduction of antioxidants glutathione and thioredoxin. TST deficiency was shown to be strongly related to the pathophysiology of metabolic diseases including diabetes and obesity. This review summarizes research related to the enzymatic properties and functions of TST, to then explore the association between the effects of TST on mitochondria and development of diseases such as diabetes and obesity.     

Unraveling the role of mitochondria during oxidative stress in plants

The sedentary habit of plants means that they must stand and fight environmental stresses that their mobile animal cousins can avoid. A range of these abiotic stresses initiate the production in plant cells of reactive oxygen and nitrogen species that ultimately lead to oxidative damage affecting the yield and quality of plant products. A complex network of enzyme systems, producing and quenching these reactive species operate in different organelles. It is the integration of these compartmented defense systems that coordinates an effective response to the various stresses. Future attempts to improve plant growth or yield must consider the complexity of inter-organelle signaling and protein targeting if they are to be successful in producing plants with resistance to a broad range of stresses. Here we highlight the role of pre-oxidant, antioxidant, and post-oxidant defense systems in plant mitochondria and the potential role of proteins targeted to both mitochondria and chloroplasts, in an integrated defense against oxidative damage in plants.     

Unraveling the regulatory role of endoplasmic-reticulum-associated degradation in tumor immunity

Abstract

During malignant transformation and cancer progression, tumor cells face both intrinsic and extrinsic stress, endoplasmic reticulum (ER) stress in particular. To survive and proliferate, tumor cells use multiple stress response pathways to mitigate ER stress, promoting disease aggression and treatment resistance. Among the stress response pathways is ER-associated degradation (ERAD), which consists of multiple components and steps working together to ensure protein quality and quantity. In addition to its established role in stress responses and tumor cell survival, ERAD has recently been shown to regulate tumor immunity. Here we summarize current knowledge on how ERAD promotes protein degradation, regulates immune cell development and function, participates in antigen presentation, exerts paradoxical roles on tumorigenesis and immunity, and thus impacts current cancer therapy. Collectively, ERAD is a critical protein homeostasis pathway intertwined with cancer development and tumor immunity. Of particular importance is the need to further unveil ERAD’s enigmatic roles in tumor immunity to develop effective targeted and combination therapy for successful treatment of cancer.     

Unraveling the role of membrane microdomains during microbial infections

Infectious diseases pose major socioeconomic and health-related threats to millions of people across the globe. Strategies to combat infectious diseases derive from our understanding of the complex interactions between the host and specific bacterial, viral, and fungal pathogens. Lipid rafts are membrane microdomains that play important role in life cycle of microbes. Interaction of microbial pathogens with host membrane rafts influences not only their initial colonization but also their spread and the induction of inflammation. Therefore, intervention strategies aimed at modulating the assembly of membrane rafts and/or regulating raft-directed signaling pathways are attractive approaches for the. management of infectious diseases. The current review discusses the latest advances in terms of techniques used to study the role of membrane microdomains in various pathological conditions and provides updated information regarding the role of membrane rafts during bacterial, viral and fungal infections.