HHV-6 has been linked in numerous studies to multiple sclerosis. Now, investigators at the University of British Columbia have published data suggesting that HHV-6A may be a key player in the development of multiple sclerosis. The investigators propose that the viral protein U24 may dysregulate myelination by strongly binding an important brain enzyme of neural precursor cell expressed developmentally down-regulated protein 4 or Nedd4.
The Canadian group were able to confirm the existence of a cognate ligand of the HHV-6A and HHV-7 U24. The ligand is called hNedd4L-WW3* domain, and it is a member of the Nedd4 family of E3 ubiquitin ligases. In CNS, Nedd4 and Nedd4-like proteins (Nedd4L) play a crucial role in promoting dendrite outgrowth and maintaining neuronal survival. . In addition, the group demonstrated that U24 binds hNedd4L-WW3* more strongly than does Nogo-A. The investigators, led by Professor Suzana Straus, PhD, suggest that preferential binding of U24 to Nedd4 may result in direct interference of Nogo-A functions, thereby contributing to MS. They also note that U24 could potentially disrupt the normal neural environment through molecular mimicry of Nogo-A.
Dysregulation of Nedd4 levels has also been associated with several types of cancer, including glioma (Zhang 2013, He 2012) and glioblastoma multiforme (Dai 2010), which have been associated with HHV-6 infection (Crawford 2009).
The viral membrane protein U24, which is expressed early in infection, impairs T cell activation (Sullivan 2008) and blocks early endosomal recycling (Sullivan 2010), which may negatively influence the morphogenesis of the myelin sheath (Winterstein 2008).
U24 also shares an identical sequence of amino acids with human myelin basic protein (MBP), which has led to investigations into the possibility of HHV-6A contributing to MS through molecular mimicry (Leibovitch 2014). That is to say, an autoreactive immune response could be generated against MBP because it contains a sequence that is identical to the immune cells’ true target: the viral U24. Indeed, MS patients exhibit a much higher frequency of T cells reactive to both U24 and MBP than controls (Tejada-Simon 2003, Cheng 2012). Straus and co-workers have also tested this mimicry hypothesis by determining U24’s ability to be phosphorylated and its interaction with Fyn-SH3, two features which link MBP to MS. They found that although U24 can be phosphorylated at the analogous threonine position to MBP (Tait 2008) and that U24 from HHV-6A binds to Fyn-SH3 (Sang 2014), these interactions are weaker than the ones in MBP. This led the team to explore the role of a different sequence in U24, called a PY motif, which it shares with Nogo-A, a protein that appears to be involved in demyelination and that has been shown to be dysregulated in MS.
The authors compared the binding affinities of the HHV-6A-specific U24, U24-6A, and the HHV-7-specific U24-7 for Nedd4-WW domains using isothermal titration calorimetry and heteronuclear single quantum coherence nuclear magnetic resonance spectroscopy. They further characterized the binding of U24-6A by phosphorylating it to simulate a version of U24-6A that is post-translationally modified in vivo. Termed pU24-6A, this U24 peptide displayed better affinity for the WW domains than the unmodified U24-6A. Although it also showed better binding affinity for the WW domain of Nedd4 than U24-7, the affinities of both pU24-6A and U24-7 were stronger than that of Nogo-A, which suggests that these peptides could interfere with the functions of Nogo-A by binding its target enzyme more stably than it can itself. It was determined that the strong binding complex affinities of the viral proteins were a result of hydrophobic contacts and hydrogen bonding, as well as the formation of salt bridges to some extent, but not of electrostatic interactions. Moreover, it was found that the interaction was not only dependent on the presence of the PY motif, but on the residues upstream from this motif.
Straus commented: “ We are excited about these new research findings, as they help us better understand the function of U24 and its potential link to MS. Going forward, we are particularly keen on seeing what the implications of this U24/WW3 domain interaction are, in particular since there are clues that analogous proteins in EBV and VZV might bind to WW3 domains in the same way.”