Despite the omnipresence of specific host–symbiont associations with acquisition of the microbial symbiont from the environment, little is known about how the specificity of the interaction evolved and is maintained. The bean bug Riptortus pedestris acquires a specific bacterial symbiont of the genus Burkholderia from environmental soil and harbors it in midgut crypts. The genus Burkholderia consists of over 100 species, showing ecologically diverse lifestyles, and including serious human pathogens, plant pathogens, and nodule-forming plant mutualists, as well as insect mutualists. Through infection tests of 34 Burkholderia species and 18 taxonomically diverse bacterial species, we demonstrate here that nonsymbiotic Burkholderia and even its outgroup Pandoraea could stably colonize the gut symbiotic organ and provide beneficial effects to the bean bug when inoculated on aposymbiotic hosts. However, coinoculation revealed that the native symbiont always outcompeted the nonnative bacteria inside the gut symbiotic organ, explaining the predominance of the native Burkholderia symbiont in natural bean bug populations. Hence, the abilities for colonization and cooperation, usually thought of as specific traits of mutualists, are not unique to the native Burkholderia symbiont but, to the contrary, competitiveness inside the gut is a derived trait of the native symbiont lineage only and was thus critical in the evolution of the insect gut symbiont.

The segment of smooth muscle regulatory light chain essential for the phosphorylation dependent activation of actomyosin motor activity and the binding of myosin heavy chain was identified. The C-terminal domain of the 20-kDa light chain, which is less conserved than the rest of the polypeptide among various muscle types, was mutated by either deletion or substitution of amino acid residues and the mutant light chains were then incorporated into myosin by subunit exchange. Deletion of Lys49-Ala'66 markedly reduced the affinity of the light chain for the heavy chain, whereas the C-terminal five residues, Lys167-Asp'71, did not contribute to the binding affinity. Deletion of Lysl49-Phelsw abolished the phosphorylation-dependent activation of actomyosin ATPase activity as well as superprecipitation activity. These results suggest that the C-terminal domain ofthe regulatory light chain is critical for transmitting the change in the conformation of the regulatory light chain induced by phosphorylation at Ser'9 to the heavy chain.The actomyosin function from vertebrate smooth muscle and nonmuscle cells is regulated by phosphorylation of the 20-kDa light chain (LC20) at Ser19 by Ca2+/calmodulindependent myosin light-chain kinase (1-3). LC20 is associated with the myosin heavy chain at the head-rod junction (4, 5) placing the phosphorylation site distally to both the catalytic and actin binding sites, which are located toward the top of the myosin head (4). The question, then, is how does the phosphorylation of LC20 at the specific site (Ser19/Thr'8) interact with the distal effector sites (ATP binding and actin binding sites) so as to activate the actin-activated myosin ATPase activity. The following findings suggest that the conformation at the head-rodjunction of myosin is altered by phosphorylation, presumably by a change in the interaction between LC20 and heavy chain, and that this is an important factor for the activation mechanism. (i) Heavy meromyosin, but not myosin subfragment 1 (S-1), containing intact regulatory light chain shows phosphorylation dependence of its actin-activated ATPase activity (6). (ii) The change in the head-rod junction conformation induced by LC20 phosphorylation can be detected by changes in susceptibility to proteases (7,8), changes in flexibility (9), and changes in shape observed by electron microscopy (10,11).It is plausible that the phosphorylation alters the conformation of LC20 and that this information is communicated to the heavy chain via LC20 heavy-chain binding sites. Skeletal muscle myosin can also be phosphorylated at the homologous site of LC20, but in this case, phosphorylation does not affect the ATPase activity of actomyosin. The difference is at least partly due to the differences in these regulatory light chains. This is underscored by the observations that, whereas smooth muscle regulatory light chain restores the Ca2+

In the symbiosis of the bean bug Riptortus pedestris with Burkholderia insecticola, the bacteria occupy an exclusive niche in the insect midgut and favor insect development and reproduction. In order to understand how the symbiotic bacteria stably colonize the midgut crypts and which services they provide to the host, we compared the cytology, physiology, and transcriptomics of free-living and midgut-colonizing B. insecticola. The analyses revealed that midgut-colonizing bacteria were smaller in size and had lower DNA content, they had increased stress sensitivity, lost motility, and an altered cell surface. Transcriptomics revealed what kinds of nutrients are provided by the bean bug to the Burkholderia symbiont. Transporters and metabolic pathways of diverse sugars such as rhamnose and ribose, and sulfur compounds like sulfate and taurine were upregulated in the midgut-colonizing symbionts. Moreover, pathways enabling the assimilation of insect nitrogen wastes, i.e. allantoin and urea, were also upregulated. The data further suggested that the midgut-colonizing symbionts produced all essential amino acids and B vitamins, some of which are scarce in the soybean food of the host insect. Together, these findings suggest that the Burkholderia symbiont is fed with specific nutrients and also recycles host metabolic wastes in the insect gut, and in return, the bacterial symbiont provides the host with essential nutrients limited in the insect food, contributing to the rapid growth and enhanced reproduction of the bean bug host.

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