To our knowledge, this is the first report of the presence of anti-TMV antibodies in human serum. We also report an association between tobacco smoking and anti-TMV antibodies. Smokers had higher serum antibody responses to TMV than non-tobacco users, although non-users were also seropositive. We further discovered that anti-TMV antibody cross-reacted with the human protein TOMM40L, a mitochondrial outer membrane subunit containing a strong amino acid homology to TMV coat protein.
Specific immune responses are elicited by pathogenic antigens present in viruses, and it is generally thought that they are not elicited by plant viruses due to their inability to replicate in animals 
. Plant viruses can be routinely found in foods we eat and in human feces but they are not thought to be involved in human health or disease 
. TMV is the best characterized plant viral pathogen and almost all of its coat protein gene sequence has been shown to be antigenic 
. TMV is widely utilized as plant virus expression vectors for vaccine antigens and biopharmaceutical protein production, but limited data are available for humans regarding immune responses elicited directly by TMV virions. A report from 1968 failed to detect serum antibodies to TMV in patients with pulmonary diseases or healthy individuals using a plant virus hemagglutination test 
_ENREF_5. By using a modified Sandwich ELISA assay, our study shows that antibodies against TMV are present in serum of all subjects, including healthy smokers as well as non-smokers. These anti-TMV antibodies are mainly IgG, the most abundant pathogen-binding antibody in human serum (response level: IgG1>IgG4>IgG3). The four human IgG subclasses, IgG1 to IgG4, differ in their affinities for binding and in their abilities to induce effector responses 
. Similar to most published serum antibody reports, IgG1 was found to provide the dominant response to TMV. Smokers had a significantly higher level of serum IgG (IgG1, IgG3 and IgG4) antibodies against TMV than non-smokers, while anti-TMV IgG (IgG1, IgG3 and IgG4) levels of smokeless tobacco users did not significantly differ from those of non-smokers. The increased production of anti-TMV antibodies in tobacco smokers may reflect humoral immune abnormalities or increased exposure.
At least 13 continuous antigenic determinants (epitopes) have been mapped at the surface of TMV coat protein and their structure has been extensively studied 
, including epitopes p34–39 and p28–42 that are shown to have antigenic activity 
. In the present study, we observed a striking sequence homology (6 continuous residues) between TMV coat protein epitope p36–41 and human TOMM40L protein p60–65. TOMM40L (also known as TOMM40B) is a potential channel-forming protein implicated in the importation of protein precursors into mitochondria. It forms part of the preprotein translocase of the outer mitochondrial membrane (TOM complex) containing TOMM22, TOMM40, TOMM40L and TOMM70 and interacts with mitochondrial targeting sequences 
. In humans, a polymorphism encoding this gene complex (TOMM40) has been associated with an increased risk of developing late-onset Alzheimer disease (AD) 
, although recent results conflict with several reports 
. Kimura et al 
has reported that serum anti-TOMM40 antibodies occur twice as often in people with AD than in healthy subjects or in patients with multiple sclerosis. Given that mitochondrial dysfunction is a common observation in Alzheimer disease, Parkinson’s disease (PD) and other neurodegenerative disorders 
, the TOM complex may very well be involved in the etiology of neurodegenerative disorders.
Our data demonstrate that anti-TMV antibodies can cross-react with TOMM40L. These observations suggest molecular mimicry between TMV and human TOMM40L protein. Molecular mimicry is a concept by which autoimmune diseases may be caused by the structural similarity of antigenic determinants. For example, an immune response against a viral sequence shared by the host and the virus can evoke a tissue-specific immune response that is capable of eliciting cell and tissue destruction 
. Based on our findings, we hypothesize a role of anti-TMV antibody in TOMM40L-mediated human health and disease.
Parkinson’s disease is the second most common neurodegenerative disorder in the U.S. About 50,000 American are diagnosed with PD each year, with more than half a million Americans affected. Epidemiologic evidence has long suggested a negative association between smoking and the risk of PD. Statistics show that PD appears to affect men at a slightly higher rate than women and is less likely to affect African Americans or Asians than whites 
. Scientists have not been able to explain this apparent lower incidence in certain populations. In our study, the significant increase of serum anti-TMV IgG in smokers was only found in African Americans but not in Caucasians, pointing out a possibility that the high immune response of TMV in African American smokers may be associated with a lower incidence of PD. Mitochondrial dysfunction has been widely implicated in the pathogenesis of PD 
. The PINK1 gene products which cause autosomal recessive familial PD are localized to mitochondria in several tissues 
and a fraction of alpha-synuclein (a major player in the pathogenesis of PD) is identified in mitochondria 
. Therefore, we speculate that the infectious agent TMV can be a driver of molecular mimicry, eliciting anti-TMV antibodies cross-reacting with TOMM40L protein to alter the host’s mitochondrial function; in smokers, anti-TMV antibodies may modulate the acquired mitochondrial defects through molecular mimicry to prevent PD development.
Molecular mimicry has been linked to a number of naturally occurring human diseases such as myasthenia gravis (the acetylcholine receptor AChRα-subunit amino acids 160–167 shows cross-reactivity with a shared homologous domain on herpes simplex virus glycoprotein D p286–293) 
and multiple sclerosis (the myelin basic protein p85–99-specific T cell clones are activated by viral mimicry peptides) 
. In the case of the observed sequence homology between TMV coat protein epitope 36–41 and human TOMM40L protein p60–65, it may be that anti-TMV antibodies influence mitochondrial function to alter disease processes. Although it might be expected that antibodies to mitochondrial proteins will not have access to the intracellular space, it should be noted that antinuclear antibodies are known to play an important part in autoimmune diseases including systemic lupus erythematosis and hepatitis C 
. The alteration of mitochondrial function induced by the TMV related molecular mechanism we have discussed may be secondary to cytotoxicity (and the disruption of cellular integrity) caused by amyloid aggregates, including out-of-register Beta–sheets as recently reported by Liu et al 
. It may also be the case that anti-TMV antibodies cross-react with other proteins important in neurodegenerative disorders. The molecular mimicry process we describe here may be primarily in the CNS or may be a systemic mechanism, as alpha-synuclein is also expressed in lymphocytes 
. Furthermore, the mechanisms of molecular mimicry we discuss could take place in microglia, which are believed to play a role in PD. Impaired mitochondrial metabolism in microglia will impair neuroinflammation and modify risk. We acknowledge that the specific influence of the molecular mimicry we report on mitochondrial function is difficult to predict. It may be that abnormalities in mitochondrial processes in PD are improved by the antibody interactions. A protective role for antibodies against a plant virus (potato virus Y) in the development of Alzheimer’s disease has been proposed 
. It is also possible that the presence of anti-TMV antibodies elicits immune responses in the brain involving toll like receptors or other immune response elements involved in the pathogenesis of PD 
Our study has a number of limitations. The study’s sample size is small, with only 20 people in each group, with uneven distribution of African American subjects. The measurement of anti-TMV antibody in serum by ELISA is semi-quantitative, and we were unable to quantitatively measure antibody levels. The study population included relatively young subjects and aged adults are at higher risk for PD, so we are unable to draw a firm conclusion regarding PD. Further studies are needed to evaluate the relationships among anti-TMV and anti-TOMM40L antibodies and the development of PD, and to determine the role of TOMM40L in pathogenesis of PD. Future study of the effects of TMV exposure on human health and disease may provide a possible therapeutic strategy against Parkinson’s disease. Finally, TMV is being used as a vector for antigen delivery in bioengineered vaccines 
. Our data suggests that immune responses to TMV antigens deserve investigation because of their potential to interfere with vaccine biodistribution or cellular metabolism.