From the mating dance of the peacock spider to brood parasitism in the common cuckoo, behavioural ecology is a fascinating and complex science. It is defined as the study of the evolutionary behaviour of animals due to ecological selection pressures, and even the smallest of organisms such as bacteria can effectively emulate eukaryotic social behaviours.
Streptomycetes are one such genus which can form multi-cellular colonies with distinct multi-nucleated hyphae structures. These hyphae have distinct compartments separated through infrequent cross-walls. The group is also significant due to their medicinal purpose; they produce over half of the world’s antibacterial and antiparasitic drugs and are commonly known for their forest like smell caused by the organic compound Geosmin.
The group harbour perplexing traits and behaviours. When mechanically macerated, the hyphae surprisingly do not ‘bleed’ to death suggesting the end is plugged and compartmentalised. Furthermore, growing hyphal tips can form up to 100 septa and in such multi-nucleated species, which lack DNA damage control proteins, we are left wondering how DNA can be protected from intense intra-cellular movement.
Imaging cells is a great way of gaining insight into the complex intracellular machinery and dynamic processes. Cryo electron microscopy is a popular method, however the process is sensitive and labour intensive. Fluorescence microscopy although sensitive to biological processes, lacks imaging of the cell morphology and resolution.
Cryo-CLEM is a technique which combines the high resolving power of electron microscopy with the specific labelling capacities of fluorescence microscopy, creating a better picture of cellular mechanics by imaging the same sample location and superimposing the complementing information.
For such a technique Linkam, alongside Professor Bram Koster and his laboratory at Leiden University Medical Centre, created the cryo-stage CMS196M. Fluorescently labelled samples are embedded in vitrified “glass like” ice and under cryo-conditions the fluorescence signal and the structural details are preserved.
September’s Paper of the Month is a collaboration between the laboratory of Professor Gilles Van Wezel and the Koster laboratory and their work using the CMS196M to answer the questions surrounding the complex behaviour of Streptomyces albus.
They conducted fluorescent light microscopy which highlighted gatherings of lipid blebs within hyphae structures. Through the use of cryo-CLEM they were able to directly label these molecules and visualise them in frozen samples allowing the study of fine cellular details in 3D.
These assemblies were termed cross-membranes and were found to span across various hyphal regions between cell wall and cell membrane.
Visualisation studies and permeability assays determined a number of these cross-membranes are impermeable, thus explaining how the bacteria may survive physical maceration. Furthermore comparative visualisation with chromosomes showed the cross-membranes create nuceloid free zones, thus explaining how the bacteria undergo severe compartmentalisation without suffering from DNA damage.
When discussing the motivations of their work, Dr Roman Koning said: “The motivation of our correlative cryo light and electron microscopy research was twofold, having both a technical and biological goal.
In terms of the biology, we found occasional intricate lipid structures in Streptomyces bacteria. Streptomyces are long bacteria that grow in soil and waters, do not divide by fission and have a branched mycelium. We wanted to be able to image specific lipid structures in Streptomyces in order to find out what their structure and function could be.
Since these structures were sparse they were difficult to find by cryo-EM and therefore we tried to locate these lipids by chemically tagging them and localizing them using fluorescence microscopy, after which we imaged the same positions using cryo electron tomography in order to get to know more of what they look like. Additionally many additional (light microscopy) experiments were performed to determine and prove what their function is.
It appeared that the lipid structures are formed by lipid tubes between the cytoplasmic membrane and the cell wall that can constrict the cell cytoplasm and thereby completely divide it, forming different cellular compartments without fission”.
Through the power of cryo-CLEM the group were able to uncover many of the perplexing mysteries surrounding the unique behaviour of Streptomyces albus. It is exciting to consider the future use of the cryo-CLEM technique and the scientific unknowns we have yet to uncover.
By Tabassum Mujtaba
Celler, K. et al. Cross-membranes orchestrate compartmentalization and morphogenesis in Streptomyces. Nat. Commun. 7:11836 doi: 10.1038/ncomms11836 (2016).