Type III Hyperlipoproteinaemia

Eighteen patients with type III hyperlipoproteinaemia, diagnosed on the basis of skin lesions, serum lipids, and lipoprotein electrophoresis, have been fully investigated over a period of 15 years. The incidence of coronary artery disease was only slightly increased, and was not increased at all among first-degree relatives. Peripheral occlusive arterial disease was probably more common. An increased incidence of carbohydrate intolerance was found in neither the patients nor their relatives. The effects of treatment on the skin were uniformly good.     

Glycogenosis Type III in the Dog

Enzyme and glycogen structure studies have been carried out on tissues of a glycogenotic dog, the clinical and pathological characteristics of which are reported in the accompanying paper. Liver glucose-6-phosphatase, leukocyte and liver acid maltase, and liver and skeletal muscle glycogen Phosphorylase all appeared largely unaffected. The activity of the muscle and liver debranching enzyme (amylo-l,6-glucosidase), determined by two independent assay methods, was, however, reduced to between 0 and 7 % of normal activity. Glycogen structure studies with Phosphorylase or iodine spectra revealed that the abnormally large amounts of glycogen found in liver and skeletal muscle had abnormally short branches, as would be expected for a deficiency of debranching enzyme. It is thus clear that the dog had suffered from the equivalent of Cori's disease (limit dextrinosis, type III glycogen storage disease). Preliminary data indicate that it may be possible to identify heterozygotes based on a study of the debranching enzyme of leukocytes.     

Immunohistochemical Video-microdensitometry of Myocardial Collagen Type I and Type III

Collagen is an essential part of the cardiac interstitium. Collagen subtypes, their location, total amount and the architecture of the fibrillar network are of functional importance. Architecture in terms of density of the fibrillar network is assumed to be reflected by the intensity of immunohistochemical staining of collagen. The aim of this study was to evaluate a video-based microdensitometric method for quantifying density expressed as absorbance of collagen subtypes I and III stained with an indirect immunoperoxidase method in myectomy specimens of patients with hypertrophic obstructive cardiomyopathy. Various factors influencing the immunohistochemical staining product and the technical properties of the image analysis system were investigated. Linearity between collagen concentration and the absorbance of the immunohistochemical staining product was demonstrated for collagen I using a dot-blot technique. Immunohistochemical collagen staining and density measure ment were easily reproducible. The cardiac disability of the patients was assessed according to the New York Heart Association (NYHA) criteria. There was a significant increase in collagen type I density with higher NYHA class, whereas no significant association was found for total collagen area fraction. Thus, video-based microdensitometry gives further insight into the structural remodelling of myocardial collagens and reveals their significance in the process of heart failure in hypertrophic cardiomyopathy.     

Porcine Type III RNA Polymerase III Promoters for Short Hairpin RNA Expression

Based on the highly conserved sequences of small nuclear RNA and small cytoplasmic RNA between vertebrate species, three porcine type III RNA polymerase III promoters, pY1, pY3 and pU6, were identified by using genomic DNA walking. To test the functional relationship of these sequences, the human H1 promoter of pSUPER-EGFP-l-neo vector was substituted with these three promoters to create the ppPol III-MCS vectors. The strength of each promoter was measured by its ability to derive expression of shRNA to repress expression of luciferase via RNA interference in the pig kidney epithelial cell line LLC-PK1. We determine that the ranking of promoter strength in descending order is pU6 > pY1 > pY3.     

Type III effector-mediated processes in Salmonella infection

Salmonella is one of the most successful bacterial pathogens that infect humans in both developed and developing countries. In order to cause infection, Salmonella uses type III secretion systems to inject bacterial effector proteins into host cells. In the age of antibiotic resistance, researchers have been looking for new strategies to reduce Salmonella infection. To understand infection and to analyze type III secretion as a potential therapeutic target, research has focused on identification of effectors, characterization of effector functions and how they contribute to disease. Many effector-mediated processes have been identified that contribute to infection but thus far no specific treatment has been found. In this perspective we discuss our current understanding of effector-mediated processes and discuss new techniques and approaches that may help us to find a solution to this worldwide problem.     

Herpesviruses and the Type III Interferon System

Type III interferons (IFNs) represent the most recently discovered group of IFNs. Together with type I IFNs (e.g. IFN-α/β), type III IFNs (IFN-λ) are produced as part of the innate immune response to virus infection, and elicit an anti-viral state by inducing expression of interferon stimulated genes (ISGs). It was initially thought that type I IFNs and type III IFNs perform largely redundant functions. However, it has become evident that type III IFNs particularly play a major role in antiviral protection of mucosal epithelial barriers, thereby serving an important role in the first-line defense against virus infection and invasion at contact areas with the outside world, versus the generally more broad, potent and systemic antiviral effects of type I IFNs. Herpesviruseses are large DNA viruses, which enter their host via mucosal surfaces and establish lifelong, latent infections. Despite the importance of mucosal epithelial cells in the pathogenesis of herpesviruses, our current knowledge on the interaction of herpesviruses with type III IFN is limited and largely restricted to studies on the alphaherpesvirus herpes simplex virus (HSV). This review summarizes the current understanding about the role of IFN-λ in the immune response against herpesvirus infections.     

Type III secretion gets an LcrV tip

Type III secretion is used by many Gram-negative pathogenic bacteria to inject effector proteins into eukaryotic host cells. Effector delivery requires a secretion apparatus, called an injectisome or needle complex, and the assembly of a translocation pore in a target-cell membrane. Recent work provides evidence that enlightens the view of how pore assembly might occur and of how the injectisome and the pore might be linked.     

Type III polyketide synthases in lichen mycobionts

Lichenized and non-lichenized fungi produce a wide range of secondary metabolites. So far, type I polyketide synthases (PKSs) are the suggested catalysts for the biosynthesis of lichen compounds. We were interested whether lichen mycobionts also contain type III PKSs, representing a class that was only recently discovered in fungi. With an alignment of known type III CHS-like genes we applied the CODEHOP strategy to design degenerate PCR primers. We further screened available fungal genomes for type III PKS genes and aligned these sequences for a phylogenetic analysis. Type III-like genes from lichen mycobionts are closely related to those known from non-lichenized fungi, but not to those of bacteria and/or plants. We conclude that type III PKS genes are ubiquitous in fungi. They are present in diverse unrelated lichen mycobionts, but their function in lichens is so far unclear.     

Harnessing Type I and Type III CRISPR-Cas systems for genome editing

CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated) systems are widespread in archaea and bacteria, and research on their molecular mechanisms has led to the development of genome-editing techniques based on a few Type II systems. However, there has not been any report on harnessing a Type I or Type III system for genome editing. Here, a method was developed to repurpose both CRISPR-Cas systems for genetic manipulation in Sulfolobus islandicus, a thermophilic archaeon. A novel type of genome-editing plasmid (pGE) was constructed, carrying an artificial mini-CRISPR array and a donor DNA containing a non-target sequence. Transformation of a pGE plasmid would yield two alternative fates to transformed cells: wild-type cells are to be targeted for chromosomal DNA degradation, leading to cell death, whereas those carrying the mutant gene would survive the cell killing and selectively retained as transformants. Using this strategy, different types of mutation were generated, including deletion, insertion and point mutations. We envision this method is readily applicable to different bacteria and archaea that carry an active CRISPR-Cas system of DNA interference provided the protospacer adjacent motif (PAM) of an uncharacterized PAM-dependent CRISPR-Cas system can be predicted by bioinformatic analysis.     

Type III protein secretion in Pseudomonas syringae

The type III secretion system is an essential virulence system used by many Gram-negative bacterial pathogens to deliver effector proteins into host cells. This review summarizes recent advancements in the understanding of the type III secretion system of Pseudomonas syringae, including regulation of the type III secretion genes, assembly of the Hrp pilus, secretion signals, the putative type III effectors identified to date, and their virulence action after translocation into plant cells.     

Plant Immunity Directly or Indirectly Restricts the Injection of Type III Effectors by the Pseudomonas syringae Type III Secretion System

Plants perceive microorganisms by recognizing microbial molecules known as pathogen-associated molecular patterns (PAMPs) inducing PAMP-triggered immunity (PTI) or by recognizing pathogen effectors inducing effector-triggered immunity (ETI). The hypersensitive response (HR), a programmed cell death response associated with ETI, is known to be inhibited by PTI. Here, we show that PTI-induced HR inhibition is due to direct or indirect restriction of the type III protein secretion system''s (T3SS) ability to inject type III effectors (T3Es). We found that the Pseudomonas syringae T3SS was restricted in its ability to inject a T3E-adenylate cyclase (CyaA) injection reporter into PTI-induced tobacco (Nicotiana tabacum) cells. We confirmed this restriction with a direct injection assay that monitored the in planta processing of the AvrRpt2 T3E. Virulent P. syringae strains were able to overcome a PAMP pretreatment in tobacco or Arabidopsis (Arabidopsis thaliana) and continue to inject a T3E-CyaA reporter into host cells. In contrast, ETI-inducing P. syringae strains were unable to overcome PTI-induced injection restriction. A P. syringae pv tomato DC3000 mutant lacking about one-third of its T3E inventory was less capable of injecting into PTI-induced Arabidopsis plant cells, grew poorly in planta, and did not cause disease symptoms. PTI-induced transgenic Arabidopsis expressing the T3E HopAO1 or HopF2 allowed higher amounts of the T3E-CyaA reporter to be injected into plant cells compared to wild-type plants. Our results show that PTI-induced HR inhibition is due to direct or indirect restriction of T3E injection and that T3Es can relieve this restriction by suppressing PTI.Plants come into contact with a myriad of microorganisms and rely on their innate immune systems to perceive potential microbial infections and induce immune responses. Plant innate immunity can be broadly portrayed as consisting of two major branches, distinguished primarily by their mode of microbe detection. The first branch is activated by extracellular pattern recognition receptors (Boller and Felix, 2009; Nicaise et al., 2009) that perceive broadly conserved molecules called pathogen (microbe)-associated molecular patterns (PAMPs; Medzhitov and Janeway, 1997; Ausubel, 2005). The response induced by this recognition is termed PAMP-triggered immunity (PTI; Jones and Dangl, 2006). A well-characterized example of PTI in plants is the recognition of and subsequent immune response to a small N-terminal region of bacterial flagellin by the FLS2 receptor kinase of Arabidopsis (Arabidopsis thaliana; Felix et al., 1999; Zipfel et al., 2004). Plant resistance (R) proteins activate the second branch of the plant innate immune system by recognizing specific pathogen effector proteins. The response induced by this recognition is termed effector-triggered immunity (ETI; Jones and Dangl, 2006). ETI and PTI induce similar innate immune responses, including ion fluxes, reactive oxygen species (ROS), and callose (β-1,3-glucan) deposition in the cell wall (Tsuda et al., 2008; Boller and Felix, 2009); however, ETI generally also includes the induction of a programmed cell death called the hypersensitive response (HR; Heath, 2000).The induction of ETI in response to a bacterial plant pathogen is generally due to the recognition of bacterial type III effector (T3E) proteins injected into the plant cell by the pathogen''s type III protein secretion system (T3SS; Alfano and Collmer, 1997; Buttner and He, 2009). These recognized T3Es were classically known as avirulence (Avr) proteins because they induced ETI responses sufficient to prevent a normally virulent pathogen from causing disease, thereby rendering it avirulent (Leach and White, 1996). However, it has become increasingly apparent that many T3Es benefit their bacteria by suppressing PTI and ETI (Block et al., 2008; Cui et al., 2009; Guo et al., 2009). Under the current model, plants first developed PTI to reduce microbial colonization of the apoplast. Successful bacterial pathogens countered this by acquiring a T3SS and PTI-suppressing T3Es (Espinosa and Alfano, 2004; Chisholm et al., 2006; Jones and Dangl, 2006).The bacterial pathogen Pseudomonas syringae infects the aerial parts of many plant species. It displays host specificity, and its strains have been separated into more than 50 pathovars based on the host plants that they infect. For example, P. syringae pv tabaci is virulent in tobacco (Nicotiana tabacum), but it triggers nonhost resistance in Arabidopsis, a plant-microbe interaction referred to as a nonhost interaction. Nonhost resistance describes the resistance observed when all members of a plant species are resistant to a specific pathogen (Thordal-Christensen, 2003; Mysore and Ryu, 2004). While not well understood, both PTI (Li et al., 2005) and ETI (Nissan et al., 2006; Wei et al., 2007) have been shown to play a role in nonhost resistance to bacterial pathogens. In some cases, P. syringae strains display race cultivar resistance. This is generally due to the resistant cultivar possessing an R protein that can recognize a T3E from the pathogen inducing ETI (Bent and Mackey, 2007). One well-studied P. syringae strain is P. syringae pv tomato DC3000, which causes bacterial speck disease on specific tomato (Solanum lycopersicum) cultivars and disease on all ecotypes of Arabidopsis tested. These interactions have been classically referred to as compatible interactions. However, DC3000 triggers nonhost resistance in tobacco and many other plants.DC3000 contains more than 30 T3Es (Lindeberg et al., 2006; Cui et al., 2009; Cunnac et al., 2009). These are encoded by genes contained within the Hrp pathogenicity island, which also encodes the T3SS apparatus (Alfano et al., 2000), other pathogenicity islands, or as individual genes throughout the genome of DC3000 (Buell et al., 2003; Wei et al., 2007). One molecular tool that has been useful in studying the effect individual T3Es have on plants is the cosmid pHIR11 (Huang et al., 1988). This cosmid encodes a functional T3SS from P. syringae pv syringae 61 and the T3E HopA1. It confers upon nonpathogenic bacteria, such as Pseudomonas fluorescens, the ability to inject HopA1 into plant cells. In tobacco and other plants, injected HopA1 induces ETI, including an HR (Huang et al., 1988; Alfano et al., 1997). The expression of other T3Es in P. fluorescens(pHIR11) enabled them to be screened for the ability to suppress HopA1-induced ETI (Jamir et al., 2004; Guo et al., 2009). Bacterial strains carrying the pHIR11 derivatives pLN18 or pLN1965, both of which lack hopA1 and so no longer induce ETI, were used to determine which T3Es could suppress PTI (Oh and Collmer, 2005; Guo et al., 2009). Collectively, these experiments demonstrated that many P. syringae T3Es possessed the ability to suppress both ETI and PTI.One PTI suppression assay using P. fluorescens(pLN18) employed by Oh and Collmer (2005) took advantage of earlier observations indicating that PTI could inhibit the ability of the plant to mount an HR in response to an ETI-inducing bacterial strain (Newman et al., 2000; Klement et al., 2003). In this assay, the PTI inducers P. fluorescens(pLN18) or a 22-amino-acid peptide from flagellin (flg22) are infiltrated into Nicotiana benthamiana. Six hours later, the ETI inducer DC3000 is infiltrated in a region of the leaf that overlaps with the earlier infiltration. The HR is typically inhibited in the overlapping region that was pretreated with a PTI inducer. Several T3Es suppressed this inhibition when they were separately delivered at time of pretreatment (Oh and Collmer, 2005). It has been speculated that the probable mechanisms for inhibition of the HR caused by PTI include impairment of delivery of T3Es that induce the HR, modification of the events downstream of T3E recognition, or a shutdown of programmed cell death (Newman et al., 2000).Here, we show that PTI inhibits the HR on tobacco because it directly or indirectly restricts the ability of P. fluorescens(pLN1965) or DC3000 to inject T3Es based on injection (translocation) assays using T3E-adenylate cyclase (CyaA) fusions. This was confirmed using an independent injection assay that monitored the amount of the cleaved in planta form of the T3E AvrRpt2. Interestingly, this injection restriction was greatly reduced in the compatible interactions between DC3000 and Arabidopsis or between P. syringae pv tabaci 11258 and tobacco. A DC3000 mutant lacking four clusters of T3E genes, which corresponds to 11 T3Es, was less able to inject a T3E-CyaA fusion into PTI-induced Arabidopsis, suggesting that the PTI suppressing activities of the T3E inventory of DC3000 allow it to overcome the injection restriction. Transgenic Arabidopsis plants separately expressing specific T3Es known to be capable of PTI suppression increased the ability of P. fluorescens(pLN1965) to inject a T3E-CyaA fusion into PTI-induced plant cells. Collectively, these data suggest that PTI can directly or indirectly restrict type III injection and PTI suppression by T3Es can relieve this restriction in susceptible plant cells but not plant cells undergoing ETI.     

III 型干扰素的抗肿瘤作用*

Ⅲ型干扰素(IFNλs)是干扰素家族中最新的一组成员。他们的很多生物学活性与临床上应用广泛的IFNα/β十分相似,但是他们所结合的受体与IFNα/β迥异。Ⅲ型干扰素的毒副作用显著小于IFNα/β,且抗肿瘤作用明显。本文回顾了近年来对Ⅲ型干扰素抗肿瘤功能研究的最新进展。     

Type III secretion: the bacteria-eukaryotic cell express

Type III secretion (T3S) is an export pathway used by Gram-negative pathogenic bacteria to inject bacterial proteins into the cytosol of eukaryotic host cells. This pathway is characterized by (i) a secretion nanomachine related to the bacterial flagellum, but usually topped by a stiff needle-like structure; (ii) the assembly in the eukaryotic cell membrane of a translocation pore formed by T3S substrates; (iii) a non-cleavable N-terminal secretion signal; (iv) T3S chaperones, assisting the secretion of some substrates; (v) a control mechanism ensuring protein delivery at the right place and time. Here, we review these different aspects focusing in open questions that promise exciting findings in the near future.     

Conformational selection and collagenolysis in Type III collagen

Matrix metalloproteases (MMPs) cleave native collagen at a single site despite the fact that collagen contains more than one scissile bond that can, in principle, be cleaved. For peptide bond hydrolysis to occur at one specific site, MMPs must (1) localize to a region near the unique scissile bond, (2) bind residues at the catalytic site that form the scissile bond, and (3) hydrolyze the corresponding peptide bond. Prior studies suggest that for some types of collagen, binding of noncatalytic MMP domains to amino acid sequences in the vicinity of the true cleavage site facilitates the localization of collagenases. In the present study, our goal was to determine whether binding to the catalytic site also plays a role in determining MMP specificity. To investigate this, we computed the conformational free energy landscape of Type III collagen at each potential cleavage site. The free energy profiles suggest that although all potential cleavage sites sample unfolded states at relatively low temperatures, the true cleavage site samples structures that are complementary to the catalytic site. By contrast, potential cleavage sites that are not cleaved sample states that are relatively incompatible with the MMP active site. Furthermore, our findings point to a specific role for arginine residues in modulating the structural stability of collagen near the collagenase cleavage site. These data imply that locally unfolded potential cleavage sites in Type III collagen sample distinct unfolded ensembles, and that the region about the true collagenase cleavage site samples states that are most complementary to the MMP active site. Proteins 2010. © 2009 Wiley‐Liss, Inc.