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Identification of hematopoietic-specific regulatory elements from the CD45 gene and use for lentiviral tracking of transplanted cells.

Abstract：

:The development of a hematopoietic reporter is crucial for determining the fate of lineages derived from cell-based therapies. A marking system will enable safer embryonic stem and induced pluripotent stem cell-based derivation of blood lineages and facilitate the development of efficient cellular reprogramming strategies based on direct fibroblast conversion. Here we report that the protein tyrosine phosphatase CD45 is an ideal candidate gene on which to base a hematopoietic reporter. CD45 regulatory elements were discovered by analyzing transcription factor chromatin occupancy (ChIP-seq) and promoter nuclease sensitivity (DNase-seq) to identify minimally sufficient sequences required for expression. After cloning the CD45 regulatory elements into an attenuated lentiviral backbone, we found that two transcriptional initiation regions were essential for high-level expression. Expressing CD45 promoters containing these regions and tethered to green fluorescent protein (GFP) in a primary B-cell differentiation assay and a transplantation model resulted in high levels of GFP in lymphoid, myeloid, and nucleated erythroid cells in mouse and human blood cell lineages. Moreover, GFP levels remained high 5 months after secondary transplantation, indicating persistence of the reporter. No CD45-driven GFP expression is observed after fibroblast or embryonic stem cell transduction. The GFP reporter is seen only after embryonic stem cells differentiate into hematopoietic cell progenitors and lineages, suggesting that this hematopoietic reporter system could be useful in validating potential autologous blood cell therapies.

journal_name

Exp Hematol

journal_title

Experimental hematology

authors

Duong KL,Das S,Yu S,Barr JY,Jena S,Kim E,Zavazava N,Colgan JD,Xue HH,Levasseur DN

doi

10.1016/j.exphem.2014.05.005

subject

Has Abstract

pub_date

2014-09-01 00:00:00

pages

761-72.e1-10

issue

9

eissn

0301-472X

issn

1873-2399

pii

S0301-472X(14)00196-9

journal_volume

42

pub_type

杂志文章

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