Modulation of Brucella-induced macropinocytosis by lipid rafts mediates intracellular replication

Watarai, Masahisa; Makino, Sou Ichi; Fujii, Y.; Okamoto, Keinosuke; Shirahata, Toshikazu

doi:10.1046/j.1462-5822.2002.00195.x

Cited by 159 publications

(153 citation statements)

References 57 publications

(67 reference statements)

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Section: Introductionmentioning

confidence: 99%

“…Neither virulence factors that allow intracellular survival of Brucella species nor the specific factors that induce such virulence have been elucidated. B. abortus is internalized by macrophages by swimming on the cell surface with generalized membrane ruffling for several minutes, a process termed 'swimming internalization', after which the bacteria are enclosed by macropinosomes (Watarai et al, 2002a).Recently we showed that Hsp60, a member of the GroEL family of chaperonins, of B. abortus is secreted on the bacterial surface through a type IV system and can interact with the cellular prion protein (PrP G ) on macrophages (Watarai et al, 2003 vacuoles rapidly fuse with early autophagosomes that acquire vacuolar H + -ATPase and lysosome-associated membrane proteins (LAMPs), mature into late autophagosomes, inhibit fusion with lysosomes and finally become replicating vacuoles, normally associated with the endoplasmic reticulum (Comerci et al, 2001; Pizarro-Cerda et al, 1998a, b). The genetic basis of B.abortus virulence is still poorly understood.…”

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Roles of Brucella abortus SpoT in morphological differentiation and intramacrophagic replication

et al. 2005

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The essential mechanisms and virulence factors enabling Brucella species to survive and replicate inside host macrophages are not fully understood. The authors previously reported that a putative guanosine 59-diphosphate 39-diphosphate (ppGpp) mutant (spoT mutant) of Brucella abortus failed to replicate in HeLa cells. The present study showed that the pattern of surface proteins and morphological change of the spoT mutant were different from B. abortus wild-type. B. abortus wild-type changed its morphology upon treatment with ppGpp synthetase I activation inhibitor. In various tests under stress conditions, including nutrient starvation, nitric oxide resistance, acid resistance and antibiotic resistance, the spoT mutant had a lower stress resistance than B. abortus wild-type. Although the spoT mutant has the same smooth phenotype and LPS profile as B. abortus wild-type, it had a higher rate of adherence to macrophages but lower internalization and intracellular replication within macrophages. The spoT mutant did not co-localize with either late endosomes or lysosomes and was almost cleared from the spleens of mice after 10 days, without splenomegaly. RT-PCR was used to detect spoT mRNA from around 10 6 cells incubated in low-pH enriched medium; it showed that the expression of spoT increased after 30 min incubation. The data suggest that SpoT does not contribute to intracellular trafficking of B. abortus, but contributes to the maintenance of bacterial morphology and the physiological adaptation required for intracellular replication. INTRODUCTIONBrucellosis is a major bacterial zoonosis and an important cause of a serious debilitating disease in humans and abortion and sterility in domestic animals. This disease is caused by species of the genus Brucella, classified in the a2 subdivision of Proteobacteria, small Gram-negative and facultative intracellular pathogens that can multiply within professional and non-professional phagocytes (Delrue et al., 2001;Detileux et al., 1990). The genus Brucella consists of seven species according to antigenic variation and primary host: B. melitensis (sheep and goats), B. suis (hogs), B. abortus (cattle), B. ovis (sheep), B. canis (dogs), B. neotomae (wood rats) and B. maris (marine mammals) (Ko & Splitter, 2003). In contrast to other intracellular pathogens, Brucella species do not produce exotoxins, antiphagocytic capsules, thick cell walls, resistance forms or fimbriae and do not show antigenic variation (Finlay & Falkow, 1997).A key aspect of the virulence of Brucella species is their ability to proliferate within professional and nonprofessional phagocytic host cells, therefore successfully bypassing the bactericidal effects of phagocytes. Their virulence and chronic infections are thought to be due to their ability to avoid the killing mechanisms within host cells (Comerci et al., 2001;Ugalde, 1999). Neither virulence factors that allow intracellular survival of Brucella species nor the specific factors that induce such virulence have been elucidated. B. abortus is inte...

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Section: Introductionmentioning

confidence: 99%

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confidence: 99%

Roles of Brucella abortus SpoT in morphological differentiation and intramacrophagic replication

et al. 2005

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“…Immature dendritic cells and macrophages both continuously take up their surrounding fluid by macropinocytosis to sample their environment and capture antigens for presentation (8). Unfortunately this process can provide a way for pathogens to gain entry into host cells and is thus exploited by a number of infectious agents such as viruses, bacteria, and even prions (9)(10)(11)(12)(13).…”

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confidence: 99%

WASH drives early recycling from macropinosomes and phagosomes to maintain surface phagocytic receptors

Buckley

Gopaldass

Bosmani

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Macropinocytosis is an ancient mechanism that allows cells to harvest nutrients from extracellular media, which also allows immune cells to sample antigens from their surroundings. During macropinosome formation, bulk plasma membrane is internalized with all its integral proteins. It is vital for cells to salvage these proteins before degradation, but the mechanisms for sorting them are not known. Here we describe the evolutionarily conserved recruitment of the WASH (WASP and SCAR homolog) complex to both macropinosomes and phagosomes within a minute of internalization. Using Dictyostelium, we demonstrate that WASH drives protein sorting and recycling from macropinosomes and is thus essential to maintain surface receptor levels and sustain phagocytosis. WASH functionally interacts with the retromer complex at both early and late phases of macropinosome maturation, but mediates recycling via retromer-dependent and -independent pathways. WASH mutants consequently have decreased membrane levels of integrins and other surface proteins. This study reveals an important pathway enabling cells to sustain macropinocytosis without bulk degradation of plasma membrane components.phagocytosis | macropinocytosis | WASH | Dictyostelium | trafficking

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“…This operon is composed of 13 open reading frames (ORFs) that share homology with other bacterial type IV secretion systems in the intracellular trafficking of pathogens. Deletion or polar and non-polar mutations of these ORFs were not able to replicate and survive within phagocytes [19,21], and internalization and uptake pathway in phagosome trafficking between wildtype and virB mutant showed different patterns [11]. Thus, the VirB proteins of B. abortus are thought to be constituents of the secretion apparatus.…”

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confidence: 99%

Zinc Uptake System (znuA Locus) of Brucella abortus is Essential for Intracellular Survival and Virulence in Mice

Kim

Watanabe

Shirahata

et al. 2004

The Journal of Veterinary Medical Science

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ABSTRACT. Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in professional and nonprofessional phagocytes, and cause abortion in domestic animals and undulant fever in humans. The mechanism and factors of virulence are not fully understood. High-affinity zinc uptake system protein mutant (znuA mutant) showed reduced growth in zinc chelated medium, and failed to replicate in HeLa cells and mouse bone marrow-derived macrophages. Transformation of znuA mutant with a shuttle vector pBBR1MCS-4 containing znuA gene restored the growth in zinc chelated medium and intracellular replication in HeLa cells and macrophages to a level comparable to that of wild-type strain. Bacterial internalization into HeLa cells and macrophages and co-localization with either late endosomes or lysosomes of znuA mutant were not different from those of wild-type strain. These results suggest that znuA does not contribute to intracellular trafficking of B. abortus, but contributes to utilization of zinc required for intracellular growth. KEY WORDS: Brucella abortus, intracellular growth, zinc, znuA.J. Vet. Med. Sci. 66(9): 1059-1063, 2004 Brucella abortus, a facultative, gram-negative, intracellular pathogen, is the etiologic agent of brucellosis, a widely distributed zoonosis [22]. The establishment of chronic infection depends on the ability of brucellae to survive within phagocytes [9]. In contrast to other intracellular pathogens, Brucella species do not produce exotoxins, antiphagocytic capsules or thick cell walls, resistance forms or fimbriae and do not show antigenic variation [7]. Intracellular survival and replication are key virulence features of Brucella because mutants defective for these attributes are also avirulent [10,14]. A key aspect of the virulence of Brucella successfully bypasses the bactericidal effects of phagocytes, and their virulence and chronic infections are thought to be due to their ability to avoid the killing mechanisms within host cells [5,20].The genetic basis of Brucella virulence is still poorly understood. The VirB type IV secretion system of Brucella has been identified recently [16]. This operon is composed of 13 open reading frames (ORFs) that share homology with other bacterial type IV secretion systems in the intracellular trafficking of pathogens. Deletion or polar and non-polar mutations of these ORFs were not able to replicate and survive within phagocytes [19,21], and internalization and uptake pathway in phagosome trafficking between wildtype and virB mutant showed different patterns [11]. Thus, the VirB proteins of B. abortus are thought to be constituents of the secretion apparatus. The genus Brucella does not contain plasmids naturally, and it is therefore possible that the proteins encoded by virB genes are involved in protein secretion rather than conjugation. A possible role in virulence is to inject effector molecules, which help in the establishment of replication niche into the host cell.Zinc plays an important role in living organism...

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Modulation of Brucella-induced macropinocytosis by lipid rafts mediates intracellular replication

Cited by 159 publications

References 57 publications

Roles of Brucella abortus SpoT in morphological differentiation and intramacrophagic replication

Roles of Brucella abortus SpoT in morphological differentiation and intramacrophagic replication

WASH drives early recycling from macropinosomes and phagosomes to maintain surface phagocytic receptors

Zinc Uptake System (znuA Locus) of Brucella abortus is Essential for Intracellular Survival and Virulence in Mice

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