抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Cell membranes display a tremendous complexity of lipids and proteins designed to perform the functions cells require. To coordinate these functions, the membrane is able to laterally segregate its constituents. This capability is based on dynamic liquid-liquid immiscibility and underlies the raft concept of membrane subcompartmentalization. Lipid rafts are fluctuating nanoscale assemblies of sphingolipid, cholesterol, and proteins that can be stabilized to coalesce, forming platforms that function in membrane signaling and trafficking. Here we review the evidence for how this principle combines the potential for sphingolipid-cholesterol self-assembly with protein specificity to selectively focus membrane bioactivity.

/pdf/lipid-rafts-as-a-membrane-organizing-principle-3mxewaemed.pdf

Lipid Rafts As a Membrane-Organizing Principle

This ebook is the first authorized digital version of Kernighan and Ritchie's 1988 classic, The C Programming Language (2nd Ed.). One of the best-selling programming books published in the last fifty years, "K&R" has been called everything from the "bible" to "a landmark in computer science" and it has influenced generations of programmers. Available now for all leading ebook platforms, this concise and beautifully written text is a "must-have" reference for every serious programmers digital library.

As modestly described by the authors in the Preface to the First Edition, this "is not an introductory programming manual; it assumes some familiarity with basic programming concepts like variables, assignment statements, loops, and functions. Nonetheless, a novice programmer should be able to read along and pick up the language, although access to a more knowledgeable colleague will help."

The C programming language

When a molecule absorbs a photon of appropriate energy, a chain of photophysical events ensues, such as internal conversion or vibrational relaxation (loss of energy in the absence of light emission), fluorescence, intersystem crossing (from singlet state to a triplet state) and phosphorescence, as shown in the Jablonski diagram for organic molecules (Fig. 1). Each of the processes occurs with a certain probability, characterized by decay rate constants (k). It can be shown that the average length of time τ for the set of molecules to decay from one state to another is reciprocally proportional to the rate of decay: τ = 1/k. This average length of time is called the mean lifetime, or simply lifetime. It can also be shown that the lifetime of a photophysical process is the time required by a population of N electronically excited molecules to be reduced by a factor of e. Correspondingly, the fluorescence lifetime is the time required by a population of excited fluorophores to decrease exponentially to N/e via the loss of energy through fluorescence and other non-radiative processes. The lifetime of photophycal processes vary significantly from tens of femotoseconds for internal conversion1,2 to nanoseconds for fluorescence and microseconds or seconds for phosphorescence.1






Open in a separate window


Figure 1


Jablonski diagram and a timescale of photophysical processes for organic molecules.

/pdf/fluorescence-lifetime-measurements-and-biological-imaging-xfalzh4w7e.pdf

Fluorescence Lifetime Measurements and Biological Imaging

The phenomenon of protein aggregation and amyloid formation has become the subject of rapidly increasing research activities across a wide range of scientific disciplines. Such activities have been stimulated by the association of amyloid deposition with a range of debilitating medical disorders, from Alzheimer's disease to type II diabetes, many of which are major threats to human health and welfare in the modern world. It has become clear, however, that the ability to form the amyloid state is more general than previously imagined, and that its study can provide unique insights into the nature of the functional forms of peptides and proteins, as well as understanding the means by which protein homeostasis can be maintained and protein metastasis avoided.

The amyloid state and its association with protein misfolding diseases

DNA point accumulation in nanoscale topography (DNA-PAINT) advances super-resolution microscopy with superior resolution and multiplexing capabilities. However, cellular DNA may interfere with this single-molecule localization technique based on DNA-DNA hybridization. Here, we introduce left-handed DNA (L-DNA) oligomers that do not hybridize to naturally present R-DNA and demonstrate that L-DNA PAINT has the same specificity and multiplexing capability as R-DNA PAINT, but greatly improves specific visualization of nuclear target molecules.

/pdf/left-handed-dna-paint-for-improved-superresolution-imaging-1fgju04qxy.pdf

Left-handed DNA-PAINT for improved superresolution imaging in the nucleus.

Septins form a filamentous collar at the mother–bud neck in budding yeast. In cytokinesis, this collar splits into two rings and the septin complexes undergo a dramatic reorientation. Using fluorescence polarization microscopy, DeMay et al. (2011. J. Cell Biol. doi:10.1083/jcb.201012143) now demonstrate that septin complexes assemble as paired filaments in vivo and reveal new insights into septin organization during cytokinesis.

/pdf/septin-pairs-a-complex-choreography-17zda345e3.pdf

Septin pairs, a complex choreography

Secondary antibodies are everyday reagents in biomedical research that are generated in animals. In this issue, Pleiner et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201709115) describe several single domain antibody fragments against antibodies from mouse and rabbit, so-called nanobodies that are easily produced recombinantly, and characterize their use in Western blotting, enzyme-linked immunosorbent assay, and immunofluorescence assays.

/pdf/open-source-recombinant-monoclonal-secondary-nanobodies-16j65k4frd.pdf

Open-source recombinant monoclonal secondary nanobodies

Septins are a conserved, essential family of GTPases that interact with actin, microtubules and membranes and form scaffolds and diffusion barriers in cells. Several of the 13 known mammalian septins assemble into nonpolar, multimeric complexes that can further polymerize into filamentous structures. While some GFP-coupled septins have been described, overexpression of GFP-tagged septins often leads to artifacts in localization and function. To overcome this ubiquitous problem, we have here generated a genome-edited rat fibroblast cell line expressing Septin 2 (Sept2) coupled to enhanced green fluorescent protein (EGFP) from both chromosomal loci. We characterize these cells by genomic polymerase chain reaction (PCR) for genomic integration, by western blot and reverse transcriptase-PCR for expression, by immunofluorescence and immunoprecipitation for the colocalization of septins with one another and cellular structures and for complex formation of different septins. By live cell imaging, proliferation and migration assays we investigate proper function of septins in these cells. We find that EGFP is incorporated into both chromosomal loci and only EGFP-coupled Sept2 is expressed in homozygous cells. We find that endogenous Sept2-EGFP exhibits expression levels, localization and incorporation into cellular septin complexes similar to the wt in these cells. The expression level of other septins is not perturbed and cell division and cell migration proceed normally. We expect our cell line to be a useful tool for the cell biology of septins, especially for quantitative biology.

https://biorxiv.org/content/biorxiv/early/2019/03/07/570622.full-text.pdf

A homozygous genome-edited Sept2-EGFP fibroblast cell line

Expansion microscopy (ExM) is a recently developed technique that allows for the resolution of structures below the diffraction limit by physically enlarging a hydrogel-embedded facsimile of the biological sample. The target structure is labeled and this label must be retained in a relative position true to the original, smaller state before expansion by linking it into the gel. However, gel formation and digestion lead to a significant loss in target-delivered label, resulting in weak signal. To overcome this problem, we have here developed an agent combining targeting, fluorescent labeling and gel linkage in a single small molecule. Similar approaches in the past have still suffered from significant loss of label. Here we show that this loss is due to insufficient surface grafting of fluorophores into the hydrogel and develop a solution by increasing the amount of target-bound monomers. Overall, we obtain a significant improvement in fluorescence signal retention and our new dye allows the resolution of nuclear pores as ring-like structures, similar to STED microscopy. We furthermore provide mechanistic insight into dye retention in ExM.

https://onlinelibrary.wiley.com/doi/pdfdirect/10.1002/anie.202302318

Helge Ewers

Papers

Left-handed DNA-PAINT for improved superresolution imaging in the nucleus.

Septin pairs, a complex choreography

Open-source recombinant monoclonal secondary nanobodies

A homozygous genome-edited Sept2-EGFP fibroblast cell line

Controlled grafting expansion microscopy.