A new intracellular pathway of haem detoxification in the midgut of the cattle tick <i>Boophilus microplus</i>: aggregation inside a specialized organelle, the hemosome

Lara, Flávio Alves; Lins, Ulysses; Paiva-Silva, Gabriela O.; Almeida, Igor C.; Braga, Claudia Mariza; Miguens, Flávio Costa; Oliveira, Pedro L.; Dansa-Petretski, Marı́lvia

doi:10.1242/jeb.00334

Cited by 107 publications

(99 citation statements)

References 27 publications

(24 reference statements)

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“…At 2·h (Fig.·5A), Pd-mP was visible as a yellow-green fluorescence, associated mainly with the large vesicles of digest cells (the vesicles implicated in hemoglobin uptake in Fig.·1A), but some fluorescence was also found in the cytosol, in contrast to hemosomes, which were almost devoid of label at this time (Fig.·5A). At 6·h (Fig.·5B), the Pd-mP fluorescence had reached the hemosomes, showing that the Pd-mP generated from digestion of Pd-mP-globin in vivo followed the same path suggested previously for heme accumulation in the digest cell (Lara et al, 2003).…”

Section: Pd-mp-globin Uptake By Digest Cells In Vivosupporting

confidence: 74%

“…Hemoglobin is thought to be hydrolyzed in the digestive vacuoles of these cells (Agbede et al, 1985;Walker and Fletcher, 1987) by the action of acid proteases (Mendiola et al, 1997), therefore releasing huge amounts of heme inside these cells. We have recently shown that most of the heme produced by this process is ultimately accumulated inside a specialized organelle called a hemosome, forming a unique type of heme aggregate (Lara et al, 2003). Using globin reconstituted with Pd-mP, a fluorescent metalloporphyrin employed here as a heme analog, we studied hemoglobin degradation in living digest cells of the cattle tick Boophilus microplus.…”

Section: Introductionmentioning

confidence: 99%

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Tracing heme in a living cell: hemoglobin degradation and heme traffic in digest cells of the cattle tickBoophilus microplus

Lara

Lins

Bechara

et al. 2005

Journal of Experimental Biology

128

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Heme is present in all cells, acting as a cofactor in essential metabolic pathways such as respiration and photosynthesis. Moreover, both heme and its degradation products, CO, iron and biliverdin, have been ascribed important signaling roles. However, limited knowledge is available on the intracellular pathways involved in the flux of heme between different cell compartments. The cattle tick Boophilus microplus ingests 100 times its own mass in blood. The digest cells of the midgut endocytose blood components and huge amounts of heme are released during hemoglobin digestion. Most of this heme is detoxified by accumulation into a specialized organelle, the hemosome.We followed the fate of hemoglobin and albumin in primary cultures of digest cells by incubation with hemoglobin and albumin labeled with rhodamine. Uptake of hemoglobin by digest cells was inhibited by unlabeled globin, suggesting the presence of receptor-mediated endocytosis. After endocytosis, hemoglobin was observed inside large digestive vesicles. Albumin was exclusively associated with a population of small acidic vesicles, and an excess of unlabeled albumin did not inhibit its uptake. The intracellular pathway of the heme moiety of hemoglobin was specifically monitored using Palladium-mesoporphyrin IX (Pd-mP) as a fluorescent heme analog. When pulse and chase experiments were performed using digest cells incubated with Pd-mP bound to globin (Pd-mP-globin), strong yellow fluorescence was found in large digestive vesicles 4·h after the pulse. By 8·h, the emission of Pd-mP was red-shifted and more evident in the cytoplasm, and at 12·h most of the fluorescence was concentrated inside the hemosomes and had turned green. After 48·h, the Pd-mP signal was exclusively found in hemosomes. In methanol, Pd-mP showed maximal emission at 550·nm, exhibiting a red-shift to 665·nm when bound to proteins in vitro.The red emission in the cytosol and at the boundary of hemosomes suggests the presence of heme-binding proteins, probably involved in transport of heme to the hemosome. The existence of an intracellular heme shuttle from the digestive vesicle to the hemosome acting as a detoxification mechanism should be regarded as a major adaptation of ticks to a blood-feeding way of life. To our knowledge, this is the first direct observation of intracellular transport of heme in a living eukaryotic cell. A similar approach, using Pd-mP fluorescence, could be applied to study heme intracellular metabolism in other cell types.

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Section: Pd-mp-globin Uptake By Digest Cells In Vivosupporting

confidence: 74%

Section: Introductionmentioning

confidence: 99%

Tracing heme in a living cell: hemoglobin degradation and heme traffic in digest cells of the cattle tickBoophilus microplus

Lara

Lins

Bechara

et al. 2005

Journal of Experimental Biology

128

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“…Whether or not this is unique to I. scapularis or that iron-binding proteins were injected at below detectable levels needs further investigation. Published evidence has suggested that the tick may detoxify heme/iron through sequestration in digestive cells (hemosomes) [118,119] and hemolymph [120][121][122]. Data in this study and others [40,41,50,52,123] that show secretion of heme binding proteins in tick saliva suggest a third possibility of eliminating heme through tick saliva.…”

Section: Heme-binding Proteinssupporting

confidence: 51%

Ixodes scapularistick saliva proteins sequentially secreted every 24h during blood feeding

Kim¹

2016

2016 International Congress of Entomology

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Ixodes scapularis is the most medically important tick species and transmits five of the 14 reportable human tick borne disease (TBD) agents in the USA. This study describes LC-MS/MS identification of 582 tick-and 83 rabbit proteins in saliva of I. scapularis ticks that fed for 24,48,72,96, and 120 h, as well as engorged but not detached (BD), and spontaneously detached (SD). The 582 tick proteins include proteases (5.7%), protease inhibitors (7.4%), unknown function proteins (22%), immunity/antimicrobial (2.6%), lipocalin (3.1%), heme/iron binding (2.6%), extracellular matrix/ cell adhesion (2.2%), oxidant metabolism/ detoxification (6%), transporter/ receptor related (3.2%), cytoskeletal (5.5%), and housekeeping-like (39.7%). Notable observations include: (i) tick saliva proteins of unknown function accounting for >33% of total protein content, (ii) 79% of proteases are metalloproteases, (iii) 13% (76/582) of proteins in this study were found in saliva of other tick species and, (iv) ticks apparently selectively inject functionally similar but unique proteins every 24 h, which we speculate is the tick's antigenic variation equivalent strategy to protect important tick feeding functions from host immune system. The host immune responses to proteins present in 24 h I. scapularis saliva will not be effective at later feeding stages. Rabbit proteins identified in our study suggest the tick's strategic use of host proteins to modulate the feeding site. Notably fibrinogen, which is central to blood clotting and wound healing, was detected in high abundance in BD and SD saliva, when the tick is preparing to terminate feeding and detach from the host. A remarkable tick adaptation is that the feeding lesion is completely healed when the tick detaches from the host. Does the tick concentrate fibrinogen at the feeding site to aide in promoting healing of the feeding lesion? Overall, these data provide broad insight into molecular mechanisms regulating different tick feeding phases. These data set the foundation for in depth I. scapularis tick feeding physiology and TBD transmission studies.PLOS Neglected Tropical Diseases |

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“…Iron from the blood meal may be in the form of heme or host-transferrinbound iron. Excessive heme resulting from hemoglobin degradation aggregates in specialized organelles called hemosomes within the digestive cells of the midgut as a detoxification mechanism (Lara et al, 2003). In the hard tick H. longicornis, intracellular ferritin has been previously identified, but its function has not been described (Xu et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

Multiple ferritins are vital to successful blood feeding and reproduction of the hard tick Haemaphysalis longicornis

Galay

Aung

Umemiya‐Shirafuji

et al. 2013

Journal of Experimental Biology

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SUMMARYTicks are obligate hematophagous parasites and important vectors of diseases. The large amount of blood they consume contains great quantities of iron, an essential but also toxic element. The function of ferritin, an iron storage protein, and iron metabolism in ticks need to be further elucidated. Here, we investigated the function a newly identified secreted ferritin from the hard tick Haemaphysalis longicornis (HlFER2), together with the previously identified intracellular ferritin (HlFER1). Recombinant ferritins, expressed in Escherichia coli, were used for anti-serum preparation and were also assayed for iron-binding activity. RT-PCR and western blot analyses of different organs and developmental stages of the tick during blood feeding were performed. The localization of ferritins in different organs was demonstrated through an indirect immunofluorescent antibody test. RNA interference (RNAi) was performed to evaluate the importance of ferritin in blood feeding and reproduction of ticks. The midgut was also examined after RNAi using light and transmission electron microscopy. RT-PCR showed differences in gene expression in some organs and developmental stages. Interestingly, only HlFER2 was detected in the ovary during oviposition and in the egg despite the low mRNA transcript. RNAi induced a reduction in post-blood meal body weight, high mortality and decreased fecundity. The expression of vitellogenin genes was affected by silencing of ferritin. Abnormalities in digestive cells, including disrupted microvilli, and alteration of digestive activity were also observed. Taken altogether, our results show that the iron storage and protective functions of ferritin are crucial to successful blood feeding and reproduction of H. longicornis. Supplementary material available online at

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A new intracellular pathway of haem detoxification in the midgut of the cattle tick Boophilus microplus: aggregation inside a specialized organelle, the hemosome

Cited by 107 publications

References 27 publications

Tracing heme in a living cell: hemoglobin degradation and heme traffic in digest cells of the cattle tickBoophilus microplus

Tracing heme in a living cell: hemoglobin degradation and heme traffic in digest cells of the cattle tickBoophilus microplus

Ixodes scapularistick saliva proteins sequentially secreted every 24h during blood feeding

Multiple ferritins are vital to successful blood feeding and reproduction of the hard tick Haemaphysalis longicornis

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