於活宿主中的全基因揭露寄生感染的新秘密＠PEREGRINE科滴

2024-08-11 20:27:51| 人123| 回0 | 上一篇

於活宿主中的全基因揭露寄生感染的新秘密

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1. 感染宿主胞的弓寄生。紫色是宿主胞及寄生的胞核，白色示寄生的外，色是由一，被肌蛋白之蛋白成的宿主胞“骨架”。

Toxoplasma gondii parasites infecting a host cell. The purple is the host cell's and parasites' nucleii, the white shows the periphery of the parasites, and the blue is the host cell's "skeleton" made of a protein called actin.

Apicomplexan parasites are a common cause of disease, infecting hundreds of millions of people each year.

寄生(一群胞的寄生)是疾病的一常原因，每年感染人。

They are responsible for spreading malaria; cryptosporidiosis – a severe childhood diarrheal disease; and toxoplasmosis – a disease that endangers immune compromised people and fetuses, and is the reason why pregnant women are told to avoid changing cat litter.

它是致播疾(孢子病，一重之童期的腹病)及弓病(一危及免疫受者胎的疾病)的原因，因此也是孕被告知，避免更砂的原因。

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Apicomplexan parasites are very good at infecting humans and many other animals, and persisting inside of them. The more that researchers can learn about how apicomplexans infect hosts, the better they will be able to develop effective treatments against the parasites.

寄生非常擅感染人及多其他物，且持存在於它。研究人能解越多，有菌如何感染宿主，越能出，此些寄生的有效法。

To this end, researchers in Whitehead Institute Member Sebastian Lourido’s lab, led by graduate student Christopher Giuliano, have now completed a genome-wide screen of the apicomplexan parasite Toxoplasma gondii (T. gondii), which causes toxoplasmosis, during its infection of mice.

此，美特海德研究所成，Sebastian Lourido室的研究人，在研究生Christopher Giuliano的下。目前，已完成有，在感染小鼠期，引弓病之寄生─弓(T. gondii)的全基因。

This screen shows how important each gene is for the parasite’s ability to infect a host, providing clues to genes’ functions. In the journal Nature Microbiology on July 8, the researchers share their approach for tracing lineages of parasites in a live host, and some specific findings of interest—including a possible anti-parasitic drug target.

示了，每一基因寄生感染宿主的能力是如何重要，而基因功能提供了多索。在(2024年)7月8日的《自然•微生物》上，此些研究人分享了，他追活宿主之寄生系的方法，及一些令人感趣的具，包括一可能之抗寄生物的的。

Researchers in Lourido’s lab previously developed a screen to test the function of every T. gondii gene in cells in a dish in 2016. They used CRISPR gene editing technology to make mutant parasites in which each lineage had one gene inactivated.

於Lourido室的研究人，先前於2016年研了一方法，培皿中，弓每一基因於胞中的功能。他使用群聚、律性隔的短文(CRISPR：Clustered Regularly Interspaced Short Palindromic Repeat)的基因技，生每一系具有一基因，遭化的突寄生。

The researchers could then assess the importance of each gene to a parasite’s fitness, or ability to thrive, based on how well the mutants missing that gene did. If a mutant died off, this implied that its inactivated gene is essential for the parasite’s survival.

然後，此些研究人能根，缺少那基因之突表怎，估每一基因寄生的健康，或快速生之能力的重要性。倘若突死亡，意味著其遭化的基因，此寄生的存活至重要。

This screen taught the researchers a lot about T. gondii’s biology but faced a common limitation: the parasites were studied in a dish rather than a live host. Cell culture provides an easier way to study parasites, but the conditions are not the same as what parasites face in an animal host. A host’s body is a more complex and dynamic environment, so it may require parasites to rely on genes that they don’t need in the artificial setting of cell culture.

使此些研究人得悉，多有弓的生物。不，面了一共同的限制：此些寄生是在培皿中，而不是在活宿主中被研究。胞培提供了一，研究寄生的方法。不，此境寄生在物宿主中，所面的境不相同。宿主的身是一，更且的境。因此，寄生可能需要仰它，在胞培之人工境中，不需要的基因。

To overcome this limitation, researchers in Lourido’s lab figured out how to repeat the T. gondii genome-wide screen, which their colleagues in the lab had previously done in cell culture, in live mice. This was a massive undertaking, which required solving various technical challenges and running a large number of parallel experiments.

了克服此限制，於Lourido室的研究人想出了，如何重弓全基因的，他於室的同僚，先前曾在活小鼠的胞培中，行。是一，需要解各技挑，行大量平行的任。

T. gondii has around eight thousand genes, so the researchers performed pooled experiments, with each mouse getting infected by many different mutants—but not so many as to overwhelm the mouse. This meant that the researchers needed a way to more closely monitor the trajectories of mutants in the mouse.

弓具有大八千基因，因而此些研究人行了多，具有每一小鼠遭多不同突感染，不不多到使小鼠受不了的集。意味著，此些研究人需要一，更密切小鼠中，多突展的方法。

They needed to track the lineages of parasites that carried the same mutation over time, as this would allow them to see how different replicate lineages of a particular mutant performed.

著推移，他需要追了相同突的寄生系。因，使他得以解，特定突的不同系如何表。

The researchers added barcodes to the CRISPR tools that inactivated a gene of interest in the parasite. When they harvested the parasites’ descendants, the barcode would identify the lineage, distinguishing replicate parasites that had been mutated in the same way.

此些研究人多添加到，化了寄生中，一重要基因的CRISPR工具中。他得此些寄生的後代，能系，分已以相同方式被突的寄生。

This allowed the researchers to use a population-based analytical approach to rule out false results and decrease experimental noise. Then they could draw conclusions about how well each lineage did. Lineage tracing allowed them to map how different populations of parasites traveled throughout the host’s body, and whether some populations grew better in one organ versus another.

使得此些研究人得以使用一，以群基的分析法，排除果及少的干。然後，他能得出有每一系，表怎的。系追使他得以，不同寄生的群，如何在整宿主的播，及某些群在一器官中，是否比在另一器官中，生得更佳。

The researchers found 237 genes that contribute to the parasite’s fitness more in a live host than in cell culture. Many of these were not previously known to be important for the parasite’s fitness. The genes identified in the current screen are active in different parts of the parasite, and affect diverse aspects of its interactions with a host.

此些研究人了237，促使此些寄生，於活宿主比在胞培中，更健康的基因。其中多先前未被知，寄生的健康是重要的。於目前中，被的此些基因，在寄生的不同部位是活的，且影其宿主交互作用的各面。

The researchers also found instances in which parasite fitness in a live host increased when a gene was inactivated; these genes may be, for example, related to signals that the host immune system uses to detect the parasites. Next, the researchers followed up on several of the fitness-improving genes that stuck out as of particular interest.

此些研究人也了多，一基因遭化，於活宿主，寄生健康提升的例。譬如，此些基因可能宿主免疫系，用此些寄生的有。接下，此些研究人行追了，在特重要，突出之改善健康的基因。

One gene that stuck out was GTP cyclohydrolase I (GCH), which codes for an enzyme involved in the production of the essential nutrient folate. Apicomplexans rely on folate, and so the researchers wanted to understand GCH’s role in securing it for the parasite.

一突出的基因是一，涉及生不可或缺素酸之酶，指定的第一型5'-三磷酸苷(GTP：Guanosine 5'-Triphosphate)水解酶(GCH)。菌依酸，因此些研究人希望解，GCH在保寄生得酸上的角色。

Cell culture media contains high levels of folate, and in this nutrient-rich environment, GCH is not essential. However, in a live mouse, the parasite must both scavenge folate and synthesize it using the metabolic pathway containing GCH. Lourido and Giuliano uncovered new details of how that pathway works.

胞培基具有高水平的酸。因此，在此富的境中，GCH非必需的。不，在活小鼠中，寄生必清除酸利用具有GCH的代途，者合成酸。Lourido及Giuliano揭露了，那途如何作的新

Although previously GCH’s role was not fully understood, the importance of folate for apicomplexans is a well-known vulnerability that has been used to design anti-parasitic therapies. The anti-folate drug pyrimethamine was commonly used to treat malaria, but many parasites have developed resistance to it.

然，先前GCH的角色不完全被解。不，酸菌的重要性是一，已被用於抗寄生法，所周知的弱。抗酸物，乙胺嘧啶(商品名拉匹林)普被用於治疾。不，多寄生已其生抗性。

Some drug-resistant apicomplexans have increased the number of GCH gene copies that they have, suggesting that they may be using GCH-mediated folate synthesis to overcome pyrimethamine. The researchers found that combining a GCH inhibitor with pyrimethamine increased the efficacy of the drug against the parasites.

一些抗性的菌已增加GCH基因的，暗示它可能利用GCH介的酸合成，抑制乙胺嘧啶。此些研究人，使GCH抑制乙胺嘧啶合，提升了此物抗此些寄生的功效。

The GCH inhibitor was also effective on its own. Unfortunately, the currently available GCH inhibitor targets mammalian as well as parasitic folate pathways, and so is not safe for use in animals. Giuliano and colleagues are working on developing a GCH inhibitor that is parasite-specific as a possible therapy.

GCH抑制其自身也有效。不巧的是，目前可用的GCH抑制，除了寄生的酸途之外，也定哺乳物。因此，供使用於物中是不安全的。Giuliano及同僚正致力於一，寄生有效之作一可能法的GCH抑制。

“There was an entire half of the folate metabolism pathway that previously looked like it wasn't important for parasites, simply because people add so much folate to cell culture media,” Giuliano says. “This is a good example of what can be missed in cell culture experiments, and what’s particularly exciting is that the finding has led us to a new drug candidate.”

Giuliano宣：「有一整半的酸代途，先前看起寄生不重要。只是因，人在胞培基中，添加了太多酸。是在多胞培中，被漏掉的一好例子。特令人振的是，此已引我朝向一新的候物。」

Another gene of interest was RASP1. The researchers determined that RASP1 is not involved in initial infection attempts, but is needed if the parasites fail and need to mount a second attempt. They found that RASP1 is needed to reload an organelle of the parasites called a rhoptry that the parasites use to breach and reprogram host cells. Without RASP1, the parasites could only deploy one set of rhoptries, and so could only attempt one invasion.

另一重要的基因是RASP1。此些研究人定了，RASP1不涉及最初的感染企。不，倘若寄生失需要第二次企，需要 RASP1。他，需要RASP1重新一，寄生用破重新宿主胞指令序列，被棒的寄生胞器。有RASP1，寄生只能一棒，因此只能企一次入侵。

Identifying the function of RASP1 in infection also demonstrated the importance of studying how parasites interact with different cell types. In cell culture, researchers typically culture parasites in fibroblasts, a connective tissue cell. The researchers found that parasites could invade fibroblasts with or without RASP1, suggesting that this cell type is easy for them to invade.

RASP1在感染中的功能也明了，研究寄生如何不同胞型交互作用的重要性。在胞培中，研究人通常，在母胞(一胞)中，培寄生。此些研究人，具有或有RASP1，寄生皆能入侵母胞。暗示，它而言，此胞型很容易入侵。

However, when the parasites tried to invade macrophages, an immune cell, those without RASP1 often failed, suggesting that macrophages present theparasites with more of a challenge, requiring multiple attempts. The screen uncovered other probable cell-type specific pathways, which would not have been found using only model cell types in a dish.

不，寄生入侵巨噬胞(一免疫胞)，那些有RASP1的往往失。暗示，巨噬胞寄生更大的挑，需要多次。此揭露了其他，於培皿中，使用模型胞型，或不曾被之可能胞型的特有途。

The screen also highlighted a previously unnamed gene that the researchers are calling GRA72. Previous studies suggested that this gene plays a role in the vacuole or protective envelope that the parasite forms around itself. The Lourido lab researchers confirmed this, and discovered additional details of how the absence of GRA72 disrupts the parasite vacuole.

此也了一，目前此些研究人GRA72，先前未被命名的基因。先前多研究暗示，基因在寄生其自身形成的液泡或保封包中，扮演一角色。Lourido室的研究人，一且了，缺少GRA72如何寄生液泡的更多。

Lourido, Giuliano, and colleagues hope that their findings will provide new insights into parasite biology and, especially in the case of GCH, lead to new therapies.

Lourido、Giuliano及同僚希望，他的研究能寄生生物，提供新的洞察力(特是有GCH)及引出新的法。

They intend to continue pulling from the treasure trove of results—their screen identified many other genes of interest that require follow-up—to learn more about apicomplexan parasites and their interactions with mammalian hosts. Lourido says that other researchers in his lab have already used the results of the screen to guide them towards relevant genes and pathways in their own projects.

他打算，多果的(他的了，多其他需要後研究的重要基因)取，得悉更多有寄生，及其哺乳物宿主的交互作用。Lourido表示，於其室的其他研究人，已使用的此些果引他，在其自己的中，朝向相的基因及途。

“This is an outstanding resource,” says Lourido, who is also an associate professor of biology at MIT. “The results of the screen reveal such a broader spectrum of ways in which the parasites are interacting with hosts, and enrich our perception of the parasites’ abilities and vulnerabilities.”