In the daylight of biotechnology:Rcpp and R applications: The Sporulation Process and Its Regulation in B. subtilis//de Hoon et al.2010

The genetically competent, non-pathogenic soil bacterium B. subtilis is the prevalent model system for studies of sporulation. A significant amount of detailed molecular data has been gathered over the years to characterize the mechanism of endospore formation — in particular, the regulation of the different sporulation stages.

Morphological Stages of Sporulation and Formation of Protective Structures

In rich medium, B. subtilis cells divide by binary fission approximately every 30 minutes. By contrast, deterioration of environmental conditions triggers sporulation, a developmental process that takes about 8 to 10 hours. Thus, endospore formation represents a formidable investment of time and energy and is considered to be a survival pathway of last resort, as B. subtilis cells only commit to sporulation after they failed to deal with starvation in other ways, such as cannibalism or establishment of a genetically competent state [14–16]. The successive morphological stages of sporulation have been defined using electron microscopy [17,18] (Figure 1). Sporulation begins with an asymmetric cell division and results in the generation of two cell types, a forespore (the smaller compartment, also called the prespore) and a mother cell. The two cells experience distinct fates, because the mother cell ultimately lyses by a programmed cell death mechanism, whereas the forespore matures as a spore. Shortly after asymmetric division, two parallel programs of gene expression are established in each compartment under the control of transcription factors that are activated in a cell-specific manner. In addition to regulatory interactions within the forespore and mother cell, precise inter-compartmental signaling is required to control the spatial and temporal progression of the developmental process. Sporulation commences only after a round of DNA replication has been completed, in order to ensure that two chromosome copies are available in the predivisional cell [19]. The two chromosomes are oriented with their origin of replication anchored at one cell pole and their origin-distal region at mid-cell [20]. After asymmetric division, only about onethird of the forespore chromosome (i.e. the origin-proximalregion) is captured in the small chamber of the dividing cell. A DNA translocase, SpoIIIE, located at the center of the polar septum, is necessary to pull the rest of this chromosome into the forespore [21–23]. The other chromosome is localized entirely inside the mother cell. Following asymmetric division, the next morphological stage of sporulation is the engulfment of the forespore by the mother cell. This process is analogous to phagocytosis and is driven by mother cell proteins that facilitate membrane migration around the forespore by enzymatic removal of the peptidoglycan [24,25]. After completion of engulfment, the forespore, now entirely surrounded by its inner and outer membranes, is a free protoplast in the mother cell cytoplasm. Next, a series of protective structures is assembled around the spore core. The cortex, a modified peptidoglycan, is synthesized between the two forespore membranes [26]. Simultaneously, at least 70 individual coat proteins are synthesized in the mother cell to encase the spore in a multi-layered structure, with the crust as the outermost layer [27,28]. Finally, the mother cell lyses to release the mature spore. Fully formed spores, recognized as the most resistant form of life on the planet [29], protect the bacterial genome against heat, desiccation, radiation, and oxidation. In addition, spore formation might be an efficient way to escape predation from higher organisms [30,31]. As soon as environmental conditions become favorable for vegetative growth, however, it is critical that B. subtilis quickly exits from the dormant state. This process is referred to as spore germination [32] and is triggered by the presence of nutrients in the environment. The nutrients are sensed by specific spore membrane receptors and, within minutes, the spore core rehydrates, the cortex is hydrolyzed, and the coat is shed. Ultimately, DNA replication is initiated and the first cell division soon follows.