Calcium-induced cleavage of DNA topoisomerase I involves the cytoplasmic-nuclear shuttling of calpain 2

Important to the function of calpains is temporal and spatial regulation of their proteolytic activity. Here, we demonstrate that cytoplasm-resident calpain 2 cleaves human nuclear topoisomerase I (hTOP1) via Ca²⁺-activated proteolysis and nucleoplasmic shuttling of proteases. This proteolysis of hTOP1 was induced by either ionomycin-caused Ca²⁺ influx or addition of Ca²⁺ in cellular extracts. Ca²⁺ failed to induce hTOP1 proteolysis in calpain 2-knockdown cells. Moreover, calpain 2 cleaved hTOP1 in vitro. Furthermore, calpain 2 entered the nucleus upon Ca²⁺ influx, and calpastatin interfered with this process. Calpain 2 cleavage sites were mapped at K¹⁵⁸ and K¹⁸³ of hTOP1. Calpain 2-truncated hTOP1 exhibited greater relaxation activity but remained able to interact with nucleolin and to form cleavable complexes. Interestingly, calpain 2 appears to be involved in ionomycin-induced protection from camptothecin-induced cytotoxicity. Thus, our data suggest that nucleocytoplasmic shuttling may serve as a novel type of regulation for calpain 2-mediated nuclear proteolysis.     

Visual perception and memory systems: from cortex to medial temporal lobe

Visual perception and memory are the most important components of vision processing in the brain. It was thought that the perceptual aspect of a visual stimulus occurs in visual cortical areas and that this serves as the substrate for the formation of visual memory in a distinct part of the brain called the medial temporal lobe. However, current evidence indicates that there is no functional separation of areas. Entire visual cortical pathways and connecting medial temporal lobe are important for both perception and visual memory. Though some aspects of this view are debated, evidence from both sides will be explored here. In this review, we will discuss the anatomical and functional architecture of the entire system and the implications of these structures in visual perception and memory.     

Beyond natural antimicrobial peptides: multimeric peptides and other peptidomimetic approaches

Naturally occurring antimicrobial peptides (AMPs) present several drawbacks that strongly limit their development into therapeutically valuable antibiotics. These include susceptibility to protease degradation and high costs of manufacture. To overcome these problems, researchers have tried to develop mimics or peptidomimetics endowed with better properties, while retaining the basic features of membrane-active natural AMPs such as cationic charge and amphipathic design. Protein epitope mimetics, multimeric (dendrimeric) peptides, oligoacyllysines, ceragenins, synthetic lipidated peptides, peptoids and other foldamers are some of the routes explored so far. The synthetic approach has led to compounds that have already entered clinical evaluation for the treatment of specific conditions, such as Staphylococcus (MRSA) infections. Should these trials be successful, an important proof-of-concept would be established, showing that synthetic oligomers rather than naturally occurring molecules could bring peptide-based antibiotics to clinical practice and the drug market for local and systemic treatment of medical conditions associated with multi-drug resistant pathogens.     

Our impasse in developing a malaria vaccine

Malaria presents a challenge to world health that to date has been beyond the abilities of researchers to conquer. This critique presents some of the strategies employed by the parasite to overcome immunity and the immunological challenges that we face to develop vaccines. A conclusion is that a vaccine must identify novel antigens or epitopes that are not normally immunogenic and which are therefore not under immune pressure and most likely to be conserved between different strains. Such antigens are most likely to be targets of cellular immunity. The case for a whole parasite blood stage vaccine is presented based on these premises.     

Evidence that prokineticin receptor 2 exists as a dimer in vivo

Prokineticins are proteins that regulate diverse biological processes including gastrointestinal motility, angiogenesis, circadian rhythm, and innate immune response. Prokineticins bind two closed related G-protein coupled receptors (GPCRs), PKR1 and PKR2. In general, these receptors act as molecular switches to relay activation to heterotrimeric G-proteins and a growing body of evidence points to the fact that GPCRs exist as homo- or heterodimers. We show here by Western-blot analysis that PKR2 has a dimeric structure in neutrophils. By heterologous expression of PKR2 in Saccharomyces cerevisiae, we examined the mechanisms of intermolecular interaction of PKR2 dimerization. The potential involvement of three types of mechanisms was investigated: coiled-coil, disulfide bridges, and hydrophobic interactions between transmembrane domains. Characterization of differently deleted or site-directed PKR2 mutants suggests that dimerization proceeds through interactions between transmembrane domains. We demonstrate that co-expressing binding-deficient and signaling-deficient forms of PKR2 can re-establish receptor functionality, possibly through a domain-swapping mechanism.     

Altered expression of securin (Pttg1) and serpina3n in the auditory system of hearing-impaired Tff3-deficient mice

Introduction

Tff3 peptide exerts important functions in cytoprotection and restitution of the gastrointestinal (GI) tract epithelia. Moreover, its presence in the rodent inner ear and involvement in the hearing process was demonstrated recently. However, its role in the auditory system still remains elusive. Our previous results showed a deterioration of hearing with age in Tff3-deficient animals.

Results

Present detailed analysis of auditory brain stem response (ABR) measurements and immunohistochemical study of selected functional proteins indicated a normal function and phenotype of the cochlea in Tff3 mutants. However, a microarray-based screening of tissue derived from the auditory central nervous system revealed an alteration of securin (Pttg1) and serpina3n expression between wild-type and Tff3 knock-out animals. This was confirmed by qRT-PCR, immunostaining and western blots.

Conclusions

We found highly down-regulated Pttg1 and up-regulated serpina3n expression as a consequence of genetically deleting Tff3 in mice, indicating a potential role of these factors during the development of presbyacusis.     

Non-conventional sources of peptides presented by MHC class I

Effectiveness of immune surveillance of intracellular viruses and bacteria depends upon a functioning antigen presentation pathway that allows infected cells to reveal the presence of an intracellular pathogen. The antigen presentation pathway uses virtually all endogenous polypeptides as a source to produce antigenic peptides that are eventually chaperoned to the cell surface by MHC class I molecules. Intriguingly, MHC I molecules present peptides encoded not only in the primary open reading frames but also those encoded in alternate reading frames. Here, we review recent studies on the generation of cryptic pMHC I. We focus on the immunological significance of cryptic pMHC I, and the novel translational mechanisms that allow production of these antigenic peptides from unconventional sources.     

Novel macrophage polarization model: from gene expression to identification of new anti-inflammatory molecules

Plasticity is a well-known property of macrophages that is controlled by different changes in environmental signals. Macrophage polarization is regarded as a spectrum of activation phenotypes adjusted from one activation extreme, the classic (M1), to the other, the alternative (M2) activation. Here we show, in vitro and in vivo, that both M1 and M2 macrophage phenotypes are tightly coupled to specific patterns of gene expression. Novel M2-associated markers were characterized and identified as genes controlling the extracellular metabolism of ATP to generate pyrophosphates (PPi). Stimulation of M1 macrophages with PPi dampens both NLR and TLR signaling and thus mediates cytokine production. In this context extracellular PPi enhanced the resolution phase of a murine peritonitis model via a decrease in pro-inflammatory cytokine production. Therefore, our study reveals an additional level of plasticity modulating the resolution of inflammation.