Intracellular traffic and fate of protein transduction domains HIV-1 TAT peptide and octaarginine. Implications for their utilization as drug delivery vectors.
Transduction domains such as those derived from the HIV-TAT protein are candidate vectors for intracellular delivery of therapeutic macromolecules such as DNA and proteins. The mechanism by which they enter cells is controversial, and very little spatial information regarding the downstream fate of these peptides from the plasma membrane is available. We studied endocytic traffic of fluorescent conjugates of HIV-TAT peptide and octaarginine in human hematopoietic cell lines K562 (CD34-) and KG1a (CD34+) and substantiated our findings in epithelia cells. Both peptides were efficiently internalized to endocytic pathways of both hematopoietic cell lines; however, comparative analysis of the intracellular location of the peptides with endocytic probes revealed major differences in spatial organization of their endocytic organelles and their interaction with the peptides at low temperatures. Double labeling confocal microscopy demonstrates that prelabeled lysosomes of all the tested cells are accessible to internalized peptides within 60 min of endocytic uptake. Incubation of cells with nocodazole and cytochalasin D inhibited peptide traffic from early to late endosomal structures, demonstrating a cytoskeletal requirement for lysosomal delivery. Disruption of Golgi and endoplasmic reticulum dynamics was without effect on peptide localization, suggesting that endosomes and lysosomes rather than these organelles are the major acceptor compartments for these molecules.
Al-Taei
Saly
S
Welsh School of Pharmacy, Cardiff University, Cardiff, CF10 3XF, Wales, United Kingdom.
Penning
Neal A
NA
Simpson
Jeremy C
JC
Futaki
Shiroh
S
Takeuchi
Toshihide
T
Nakase
Ikuhiko
I
Jones
Arwyn T
AT
D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D003911:Dextrans / Q000378:metabolism; D016503:Drug Delivery Systems; D004705:Endocytosis; D015696:Gene Products, tat / Q000378:metabolism*; D015497:HIV-1; D006367:HeLa Cells; D006801:Humans; D020014:K562 Cells; D008247:Lysosomes / Q000378:metabolism*; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D054322:tat Gene Products, Human Immunodeficiency Virus
D003911:Dextrans; D015696:Gene Products, tat; D009842:Oligopeptides; D010455:Peptides; C448619:octaarginine; D054322:tat Gene Products, Human Immunodeficiency Virus