In bacterial biotechnology, instead of producing functional proteins from plasmids, it is often necessary to deliver functional proteins directly into live cells for genetic manipulation or physiological modification. We constructed a library of cell-penetrating peptides (CPPs) capable of delivering protein cargo into bacteria and developed an efficient delivery method for CPP-conjugated proteins. We screened the library for highly efficient CPPs with no significant cytotoxicity in Escherichia coli and developed a model for predicting the penetration efficiency of a query peptide, enabling the design of new and efficient CPPs. As a proof-of-concept, we used the CPPs for plasmid curing in E. coli and marker gene excision in Methylomonas sp. DH-1. In summary, we demonstrated the utility of CPPs in bacterial engineering. The use of CPPs would facilitate bacterial biotechnology such as genetic engineering, synthetic biology, metabolic engineering, and physiology studies.

Differentiation of oligodendrocytes (ODs) presents a challenge in regenerative medicine due to their role in various neurological diseases associated with dysmyelination and demyelination. Here, we designed a peptide derived from vitronectin (VN) using in silico docking simulation and examined its use as a synthetic substrate to support the differentiation of ODs derived from human pluripotent stem cells. The designed peptide, named VNP2, promoted OD differentiation induced by the overexpression of SOX10 in OD precursor cells compared with Matrigel and full-length VN. ODs differentiated on VNP2 exhibited greater contact with axon-mimicking nanofibers than those differentiated on Matrigel. Transcriptomic analysis revealed that the genes associated with morphogenesis, cytoskeleton remodeling, and OD differentiation were upregulated in cells grown on VNP2 compared with cells grown on Matrigel. This new synthetic VN-derived peptide can be used to develop a culture environment for efficient OD differentiation.

The ability to generate astrocytes from human pluripotent stem cells (hPSCs) offers a promising cellular model to study the development and physiology of human astrocytes. The extant methods for generating functional astrocytes required long culture periods and there remained much ambiguity on whether such paradigms follow the innate developmental program. In this report, we provided an efficient and rapid method for generating physiologically functional astrocytes from hPSCs.Overexpressing the nuclear factor IB in hPSC-derived neural precursor cells induced a highly enriched astrocyte population in 2 weeks. RNA sequencing and functional analyses demonstrated progressive transcriptomic and physiological changes in the cells, resembling in vivo astrocyte development. Further analyses substantiated previous results and established the MAPK pathway necessary for astrocyte differentiation. Hence, this differentiation paradigm provides a prospective in vitro model for human astrogliogenesis studies and the pathophysiology of neurological diseases concerning astrocytes.