Molecular evidence of inefficient transduction of proliferating human B lymphocytes by VSV-pseudotyped HIV-1-derived lentivectors

M. Serafini, L. Naldini, M. Introna

Research output: Contribution to journalArticlepeer-review

Abstract

Lentiviral vectors are attractive tools to transduce dividing and nondividing cells. Human tonsillar B lymphocytes have been purified and induced to proliferate by the addition of anti-CD40 + IL-4 or anti-CD40 + anti-μ signals and transduced at high MOI with a VSV pseudotyped lentivector carrying the eGFP gene under the control of the PGK promoter. Parallel cultures of PHA-stimulated T lymphocytes containing a comparable amount of cycling cells during the infection reached over 70% eGFP transduction. By contrast, only less than 3% B lymphocytes became eGFP positive after 7 days from transduction. Molecular analysis of the viral life cycle shows that cytoplasmic retrotranscribed cDNA and nuclear 2LTR circles are detectable at lower levels and for a shorter period of time in proliferating B cells with respect to proliferating T lymphocytes. Moreover, FACS-sorted eGFP-positive and negative B cell populations were both positive for the presence of retrotranscribed cDNA and 2LTR circles nuclear forms. By contrast, nested Alu-LTR PCR allowed us to detect an integrated provirus in FACS-sorted eGFP-positive cells only. Together with the demonstration that infection in saturation conditions led to an increase in the percentage of transduced cells (reaching 9%), these findings suggest that in proliferating B lymphocytes, lentiviral transduction is an inefficient process blocked at the early steps of the viral life cycle possibly involving partially saturable restriction factors.

Original languageEnglish
Pages (from-to)413-424
Number of pages12
JournalVirology
Volume325
Issue number2
DOIs
Publication statusPublished - Aug 1 2004

Keywords

  • HIV-1
  • Lentivector
  • Lymphocyte

ASJC Scopus subject areas

  • Virology
  • Infectious Diseases

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