Revealing the inner beauty by 3D structured illumination microscopy

Posted by Hamed Shateri Najafabadi

John W. Sedat and his team at the University of California, San Francisco, have developed a revolutionary method for visualizing the cells. Their new “3D-structured illumination microscopy”, or 3D-SIM, analyzes the changes that happen in the light interference pattern when a fine cellular structure reflects the light, and interprets the image with a resolution of about 100nm, almost twice as good as the resolution of the state-of-the-art confocal laser scanning microscopes – shamefully, that was all I could understand from the physics of this microscope! This technique is fascinating in that, in contrast to the electrone microscopy techniques, it can be used for specific labeling of molecules using the very conventional methods such as labeling with fluorescent antibodies. There is no need to change the protocols that you currently use for preparing the specimens; just the micropscope is different. In a report in Science, Sedat and his team demonstrate the ability of this technique in resolving multicolor images of the nuclear periphery with an unprecedented precision, revealing exciting features such as presence of chromatin-deprived spaces just below the foci of nuclear pore complexes (see figure below).

Simultaneous imaging of DNA, nuclear lamina, and NPC epitopes by 3D-SIM

Some images are so stunning:

Cell division

A nucleus from a mouse-muscle stem cell

P.S. Also see the first comment; Marie-Luise has kindly written a description on the physics behind 3D-SIM.

Altruism in Leishmania: apoptotic parasites are required for infectivity of metacyclic promastigotes

Posted by Kasra Hassani

Suppression of the innate immune response and inhibition of activation of phagocytes that would otherwise kill the parasites has long been established as mechanisms of immune evasion and persistence among Leishmania parasites.

In their paper, van Zandbergen et al. have indicated presence of a high ratio (more than 40%) of apoptotic cells in the metacyclic/stationary phage parasites. They have characterized these cells by occurrence of phosphatidyl serine (PS) in the outer leaflet of plasma membrane as well as PS-binding protein Anexin A5(AnxA5). The majority of AnxA5+ cells have been shown to be apoptotic and different in morphology to infective parasites and they have shown that depletion of these apoptotic cells from the infective population substantially abrogates infectivity.

Apoptotic cells induce production of TGF-beta and IL-10 which are anti-inflammatory cytokines; these cytokines are produced as well by neutrophils when they phagocyte apoptotic Leishmania. Apoptotic parasites also hamper secretion of TNF-alpha, all of which results in inactivation of neutrophils and later macrophages and their inability to kill the phagocytosed parasites.

This is an interesting example of altruism among single-cell populations; the authors have suggested that apoptosis is probably triggered in late log phase and stationary phase promastigotes in the sandfly midgut due to nutrient depletion prior to their entry into the mammalian host.

Studying the secretome of Leishmania donovani

Posted by Kasra Hassani

In this paper, Silverman et al. have pointed to two interesting subjects: first, what proteins are generally secreted from Leishmania, and second, how are these proteins secreted. In an extensive proteomic analysis, they have pointed out 151 proteins that they believe are being actively secreted out of stationary promastigotes of Leishmania donovani. These proteins belong to a wide variety of groups, such as proteases, antioxidants, nucleases etc. and each might play roles in survival of the parasite within its hosts and modulation of the immune response. Identification of these proteins opens up many opportunities for further studies that promote understanding their function and possible therapeutic targets in continuing studies.

Another interesting finding of Silverman et al. was that among these secreted proteins only 2 contain a classical amino-terminal secretion signal, which means that Leishmania largely might benefit from non-classical secretion pathways such as exosomes. Exosomes have been studied previously in human B cells and dendritic cells and it is actually interesting to point out that there is striking correspondence between the proteome content of these exosomes and Leishmania’s secretome (except for the proteins for which Leishmania does not have an ortholog). The authors have proposed the release of exosomes from the surface of the cell and especially from the flagellar pocket to be an important pathway of protein secretion by Leishmania and they have observed vesicular budding from the parasite surface by STM.

microRNA processing by splicing

Posted by Hamed Shateri Najafabadi

My colleage, Yuan, sent me a paper a while ago that had interesting points on possible mechanisms of microRNA processing. Ruby, Jan and Bartel from MIT in their paper, “Intronic microRNA precursors that bypass Drosha processing” (Nature 2007, 448:83-86), report a new mechanism for maturation of microRNAs from introns. This Drosha-independent mechanism recruits the splicing machinery to produce pre-miRNAs/introns that the authors, tastefully, call ‘mirtrons’ (the conventional pathway requires the enzyme Drosha to cleave pri-miRNA transcripts into pre-miRNAs, see Figure below). This paper is a beautiful example of how high-throughput sequencing, spiced by open minds and sharp eyes can lead into spectacular findings. I would like to quote the last sentences of this paper: “This mechanism, together with that of mirtron processing, would enable miRNAs to emerge in any organism with both splicing and post-transcriptional RNA silencing, even those lacking the specialized RNase III enzyme Drosha or its plant counterpart, DICER-LIKE1. In this scenario, miRNAs might have emerged in ancient eukaryotes before the advent of modern miRNA biogenesis pathways.”

Model for convergence of the canonical and mirtronic miRNA biogenesis pathways

The Parasite Diary: an introduction

What this blog tends to do:

This blog intends to be a start-up place to bring together students and researchers from all around the world that are working on parasites that cause human diseases. We believe that much knowledge could be acquired through communication and interaction and that especially this communication is essential in the complex world of parasites.

Our goals are sharing of knowledge, ideas and technical expertise among students and researchers. The more active members our community has the more benefit we will have from one another.

How we like to start:

We like to start by putting up summaries and interesting points about research articles that we read every now and then. We also invite everyone to send us summaries of what they have found interesting and worthy of sharing and we would be pleased to post them on the blog (please see the about us page). Feel free to leave comments and initiate discussions that could go more in depth into the experimentation or data analysis.

A constitution for further development of this blog is under preparation. Ideas are more than welcome.