Inhibition of cyclin‐dependent kinase 5 but not of glycogen synthase kinase 3‐β prevents neurite retraction and tau hyperphosphorylation caused by secretable products of human T‐cell leukemia virus type I‐infected lymphocytes

Abstract

Human T-cell leukemia virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is a neurodegenerative disease characterized by selective loss of axons and myelin in the corticospinal tracts. This central axonopathy may originate from the impairment of anterograde axoplasmic transport. Previous work showed tau hyperphosphorylation at T181 in cerebrospinal fluid of HAM/TSP patients. Similar hyperphosphorylation occurs in SH-SY5Y cells incubated with supernatant from MT-2 cells (HTLV-I-infected lymphocytes secreting viral proteins, including Tax) that produce neurite shortening. Tau phosphorylation at T181 is attributable to glycogen synthase kinase 3-β (GSK3-β) and cyclin-dependent kinase 5 (CDK5) activation. Here we investigate whether neurite retraction in the SH-SY5Y model associates with concurrent changes in other tau hyperphosphorylable residues. Threonine 181 turned out to be the only tau hyperphosphorylated residue. We also evaluate the role of GSK3-β and CDK5 in this process by using specific kinase inhibitors (LiCl, TDZD-8, and roscovitine). Changes in both GSK3-β active and inactive forms were followed by measuring the regulatory phosphorylable sites (S9 and Y216, inactivating and activating phosphorylation, respectively) together with changes in β-catenin protein levels. Our results showed that LiCl and TDZD-8 were unable to prevent MT-2 supernatant-mediated neurite retraction and also that neither Y216 nor S9 phosphorylations were changed in GSK3-β. Thus, GSK3-β seems not to play a role in T181 hyperphosphorylation. On the other hand, the CDK5 involvement in tau phosphorylation was confirmed by both the increase in its enzymatic activity and the absence of MT-2 neurite retraction in the presence of roscovitine or CDK5 siRNA transfection.

Tax and Semaphorin 4D Released from Lymphocytes Infected with Human Lymphotropic Virus Type 1 and Their Effect on Neurite Growth

“…A retraction model with differentiated neuroblastoma cells (SH-SY5Y cells) gave us similar effects with culture media from both MT2 or PBMCs from HAM/TSP patients confirming the validity of using this cell line as well. 19,23 In conclusion, the interaction between secreted Tax and soluble SEMA-4D from HTLV-1-infected human lymphocytes led us to suggest a leading role for Tax in the SEMA-4D-Plexin 1B signaling pathway. The chemoattractant effect of sSEMA-4D on infected PBMCs exhibiting CRMP-2 pSer 522 implies a higher CNS infiltration of infected T cells, increasing both the occurrence of SEMA-4D T cells and sSEMA-4D levels in CSF.…”

“…22 Our group has reported that Tax secreted from an HTLV-1-infected human T cell line (MT2) produced retraction in neuroblastoma cells, SH-SY5Y. 23 This effect could be mediated by Tax as well as by other proteins released from HTLV-1-infected lymphocytes such as a proteolytically shed form of Semaphorin 4D, SEMA-4D (150-kDa transmembrane glycoprotein), called soluble Semaphorin 4D, sSEMA-4D. This soluble semaphorin is a bioactive soluble form of 120-kDa that upon binding to its neuronal receptor Plexin B1 induces growth cone collapse.…”

Human T-lymphotropic virus proteins and post-translational modification pathways

“…Valenzuela and collaborators recently found that cyclindependent kinase (Cdk)5 prevents Tau hyper-phosphorylation caused by HTLV-1-infected cell secretions [31] , but further studies are needed to understand the relevance of those modifications.…”

Tax Posttranslational Modifications and Interaction with Calreticulin in MT-2 Cells and Human Peripheral Blood Mononuclear Cells of Human T Cell Lymphotropic Virus Type-I-Associated Myelopathy/Tropical Spastic Paraparesis Patients

“…18 BDNF was removed from the culture medium and replaced by a mixture of DMEM-F12 Ham without fetal bovine serum 4 h previous to the addition of culture medium of PBMCs. In all experiments DMEM-F12 was replaced by PBMC culture medium, being differentiated in SH-SY5Y cells incubated up to 2 h with (1) culture medium of PBMCs from an HAM/TSP patient, (2) culture medium of PBMCs from an HAM/TSP patient with 10 ll/ml of either Tax antibody (HTLV-1 Tax Hyb 168A51-2) or a same isotype irrelevant antibody (anti-Gizzerosine), and (3) with culture medium of PBMCs from a healthy subject.…”

“…17 Incubation of human SH-SY5Y neuroblastoma cells with culture medium of MT-2 cells (an HTLV-1-infected cell line that secretes viral Tax protein) produces neurite retraction and an increase in Tau phosphorylation at T181. 18 Tax, a 40-kDa protein, undergoes posttranslational modifications such as phosphorylation, ubiquitination, SUMOylation, and acetylation. 15,16,[19][20][21][22][23][24] Phosphorylation is critical for Tax transactivation via both the ATF/CREB and NF-jB pathways.…”