Abstract
Homeoproteins are a class of transcription factors defined by the structure of their DNA-binding domain, the homeodomain. In addition to their nuclear cell-autonomous activities, homeoproteins transfer between cells, thanks to two separate steps of secretion and internalization, which both rely on unconventional mechanisms. Internalization is driven by the third helix of the homeodomain (Penetratin) through a non-vesicular and endocytosis-independent mechanism. In contrast, homeoprotein secretion involves vesicular compartments and requires the presence of a sequence of 11 amino acids (Sec sequence) spanning between the second and third helix of the homeodomain. In this study, we report that the SecPen polypeptide, which combines the two identified domains, Penetratin and Sec, bears all of the necessary information to go in and out of cells. We have analyzed key mechanisms and demonstrated that this peptide can efficiently cross a tight junction epithelium.
Highlights
Nalization is driven by the third helix of the homeodomain [6]
The Sec sequence, partially overlapped with Penetratin within the Engrailed homeodomain, and we investigated whether SecPen, a 27-amino-acid-long peptide that combines Penetratin and the Sec sequences, bears all of the necessary information to go in and out of cells
We demonstrated that this SecPen peptide efficiently crosses a tight junction epithelium and analyzed some key steps involved
Summary
Peptide Synthesis—Biotinylated peptides were synthesized by Neosystem (France) with a biotin residue at the N terminus using the following sequences: Pen (RQIKIWFQNRRMKWKK), PenPro (RQPKIWFPNRRMPWKK), Tat (GRKKRRQRRRP), SecPen (QSLAQELGLNERQIKIWFQNRRMKWKK), and SecTat (QSLAQELGLNERQIKIGRKKRRQRRRP). For Transwell studies, cells were seeded at 1.5 ϫ 105 cells on a
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