Báo cáo khoa học: Pleiotrophin inhibits HIV infection by binding the cell surface-expressed nucleolin

Pleiotrophin Inhibits HIV Infection By Binding the Cell Surface-Expressed Nucleolin

Tác giả: Elias A. Said, José Courty, Josette Svab, Jean Delbé, Bernard Krust, and Ara G. Hovanessian
Lĩnh vực: Vi sinh học, Y học
Nội dung tài liệu: Nghiên cứu này khám phá vai trò của yếu tố tăng trưởng pleiotrophin (PTN) trong việc ức chế sự lây nhiễm HIV. Kết quả cho thấy PTN có khả năng ngăn chặn sự gắn kết của hạt virus HIV vào bề mặt tế bào. Các miền β-sheet của PTN, đặc biệt là các axit amin từ 60-110, dường như đóng vai trò quan trọng trong cơ chế ức chế này. PTN liên kết với bề mặt tế bào thông qua các vị trí liên kết ái lực cao và ái lực thấp. Các chất ức chế gắn kết HIV khác, như HB-19, cũng liên kết với nucleolin bề mặt tế bào, cho thấy nucleolin là một thụ thể ái lực thấp cho PTN. Quan sát bằng kính hiển vi huỳnh quang cho thấy PTN tạo thành cụm trên bề mặt tế bào cùng với nucleolin. PTN được nội hóa vào tế bào thông qua một cơ chế phụ thuộc nhiệt độ, có thể bị ức chế bởi HB-19 và không phụ thuộc vào heparan và chondroitin sulfate proteoglycans. Nghiên cứu này lần đầu tiên chỉ ra rằng PTN ức chế nhiễm HIV và đề xuất nucleolin bề mặt tế bào là một thụ thể ái lực thấp cho PTN, đồng thời đóng vai trò trong quá trình nội hóa PTN vào tế bào.
Mục lục chi tiết:
– Keywords: binding; HIV; pleiotrophin; receptor; surface nucleolin
– Correspondence
– Received 11 May 2005, revised 30 June 2005, accepted 18 July 2005
– doi:10.1111/j.1742-4658.2005.04870.x
– The growth factor pleiotrophin (PTN) has been reported to bind heparan sulfate and nucleolin, two components of the cell surface implicated in the attachment of HIV-1 particles to cells. Here we show that PTN inhibits HIV-1 infection by its capacity to inhibit HIV-1 particle attachment to the surface of permissive cells. The β-sheet domains of PTN appear to be implicated in this inhibitory effect on the HIV infection, in particular the domain containing amino acids 60–110. PTN binding to the cell surface is mediated by high and low affinity binding sites. Other inhibitors of HIV attachment known to bind specifically surface expressed nucleolin, such as the pseudopeptide HB-19 and the cytokine midkine prevent the binding of PTN to its low affinity-binding site. Confocal immunofluorescence laser microscopy revealed that the cross-linking of surface-bound PTN with a specific antibody results in the clustering of cell surface-expressed nucleolin and the colocalization of both PTN and nucleolin signals. Following its binding to surface-nucleolin, PTN is internalized by a temperature sensitive mechanism, a process which is inhibited by HB-19 and is independent of heparan and chondroitin sulfate proteoglycans. Nevertheless, proteoglycans might play a role in the concentration of PTN on the cell surface for a more efficient interaction with nucleolin. Our results demonstrate for the first time that PTN inhibits HIV infection and suggest that the cell surface-expressed nucleolin is a low affinity receptor for PTN binding to cells and it is also implicated in PTN entry into cells by an active process.
– The human immunodeficiency virus (HIV) infects target cells by the capacity of its envelope glycoproteins gp120-gp41 to attach cells and induce the fusion of virus to cell membranes, a process which leads to virus entry. The receptor complex essential for HIV entry consists of the CD4 molecule and at least one of the members of the chemokine receptor family: CCR5 or CXCR4 [1,2]. Contrary to the virus entry process, the attachment of HIV particles to cells can occur even independently of CD4. We have previously demonstrated that HIV attachment is inhibited by the pseudopeptide HB-19 that binds specifically the C-terminal tail of nucleolin, a cell-surface-expressed protein identified to be implicated in HIV attachment [3-5]. Consequently, we have suggested that HIV attachment is achieved by the coordination of at least two events implicating on the one hand heparan sulfate proteoglycans [6,7] and on the other hand the cell surface-expressed nucleolin [4]. In the search for natural ligands of nucleolin that exhibit a potential inhibitory activity against HIV infection, other than midkine [8] and lactoferrin [9], here we show that pleiotrophin
– Abbreviations
– Figure 1. Inhibition of HIV infection by PTN.
– Figure 2. The inhibitory of various PTN domains on HIV infection.
– Figure 3. Attachment of HIV particles to CD4 CHO cell lines, expressing or not expressing heparan/chondroitin sulfate proteoglycans is inhibited by PTN and the nucleolin-binding HB-19 pseudopeptide.
– Figure 4. The binding of 125I-labeled PTN to CHO cell lines.
– Table 1. High affinity and low affinity Pleiotrophin receptors on CHO wild-type and proteoglycan-free cells.
– Figure 5. The effect of the nucleolin binding molecules on the binding of PTN to the low and high affinity-binding site.
– Figure 6. Internalization of PTN does not require expression of heparan (HS)/chondroitin sulfate (CS) proteoglycans.
– Figure 7. Internalization of PTN is inhibited by the HB-19 pseudopeptide.
– Figure 8. PTN induced clustering of nucleolin in MT4 cells: colocalization of PTN with nucleolin at the surface of PTN treated cells.
– Discussion
– Experimental procedures
– Materials
– Antibodies
– Cell lines and virus preparation
– HIV infection of HeLa CD4+ cells
– Expression of PTN constructs by CHO-K1 cells
– HIV attachment on HeLa CD4+ and CHO cell lines
– Assay of 125I-labeled PTN Binding to cells
– Confocal microscopy
– Acknowledgements
– References