title: Establishment of single-cell screening system for the rapid identification of transcriptional modulators involved in direct cell reprogramming.
abstract: Combinatorial interactions of transcription modulators are critical to regulate cell-specific expression and to drive direct cell reprogramming (e.g. trans-differentiation). However, the identification of key transcription modulators from myriad of candidate genes is laborious and time consuming. To rapidly identify key regulatory factors involved in direct cell reprogramming, we established a multiplex single-cell screening system using a fibroblast-to-monocyte transition model. The system implements a single-cell 'shotgun-transduction' strategy followed by nested-single-cell-polymerase chain reaction (Nesc-PCR) gene expression analysis. To demonstrate this, we simultaneously transduced 18 monocyte-enriched transcription modulators in fibroblasts followed by selection of single cells expressing monocyte-specific CD14 and HLA-DR cell-surface markers from a heterogeneous population. Highly multiplex Nesc-PCR expression analysis revealed a variety of gene combinations with a significant enrichment of SPI1 (86/86) and a novel transcriptional modulator, HCLS1 (76/86), in the CD14(+)/HLA-DR(+) single cells. We could further demonstrate the synergistic role of HCLS1 in regulating monocyte-specific gene expressions and phagocytosis in dermal fibroblasts in the presence of SPI1. This study establishes a platform for a multiplex single-cell screening of combinatorial transcription modulators to drive any direct cell reprogramming.
authors.affiliation: Omics Science Center, RIKEN Yokohama, 1-7-22 Suehiro-cho Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Jay W
authors.initials: JW
authors.lastname: Shin
authors.forename: Takahiro
authors.initials: T
authors.lastname: Suzuki
authors.forename: Noriko
authors.initials: N
authors.lastname: Ninomiya
authors.forename: Mami
authors.initials: M
authors.lastname: Kishima
authors.forename: Yuki
authors.initials: Y
authors.lastname: Hasegawa
authors.forename: Atsutaka
authors.initials: A
authors.lastname: Kubosaki
authors.forename: Haruka
authors.initials: H
authors.lastname: Yabukami
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Hayashizaki
authors.forename: Harukazu
authors.initials: H
authors.lastname: Suzuki
mesh_terms: D054338:Cell Transdifferentiation / Q000235:genetics*; D002478:Cells, Cultured; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D009000:Monocytes / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D059010:Single-Cell Analysis / Q000379:methods*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D000099232:Proto-Oncogene Protein Spi-1
chemical_list: D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; D000099232:Proto-Oncogene Protein Spi-1