Evaluating Common Humoral Responses against Fungal Infections with Yeast Protein Microarrays

Abstract

We profiled the global immunoglobulin response against fungal infection by using yeast protein microarrays. Groups of CD-1 mice were infected systemically with human fungal pathogens (Coccidioides posadasii, Candida albicans, or Paracoccidioides brasiliensis) or inoculated with PBS as a control. Another group was inoculated with heat-killed yeast (HKY) of Saccharomyces cerevisiae. After 30 days, serum from mice in the groups were collected and used to probe S. cerevisiae protein microarrays containing 4800 full-length glutathione S-transferase (GST)-fusion proteins. Antimouse IgG conjugated with Alexafluor 555 and anti-GST antibody conjugated with Alexafluor 647 were used to detect antibody-antigen interactions and the presence of GST-fusion proteins, respectively. Serum after infection with C. albicans reacted with 121 proteins: C. posadasii, 81; P. brasiliensis, 67; and after HKY, 63 proteins on the yeast protein microarray, respectively. We identified a set of 16 antigenic proteins that were shared across the three fungal pathogens. These include retrotransposon capsid proteins, heat shock proteins, and mitochondrial proteins. Five of these proteins were identified in our previous study of fungal cell wall by mass spectrometry (Ann. N. Y. Acad. Sci. 2012, 1273, 44-51). The results obtained give a comprehensive view of the immunological responses to fungal infections at the proteomic level. They also offer insight into immunoreactive protein commonality among several fungal pathogens and provide a basis for a panfungal vaccine.

Proteomic Analysis of Pathogenic Fungi Reveals Highly Expressed Conserved Cell Wall Proteins

“…Using studies in vitro , the chance of this can be maximized by using proteins that are expressed in multiple growth conditions. Such expression would likely also be conserved in vivo , as supported by other published observations [ 61 , 88 ]. A superior vaccine may be created by selecting immunogenic epitopes of several proteins and combining them into a recombinant protein.…”

“…In recent studies, we tested serum from mice infected with C. posadasii , C. albicans or Paracoccidioides brasiliensis on a 4800 element ordered protein array of Saccharomyces cerevisiae to profile cross-reactive antigen proteins [ 88 ]. These sera detected 60 to 130 proteins each, including five that we had reported in our initial studies using immunoblotting and MS [ 61 ]; 16 detected proteins were common to all.…”

Role of Heat-Shock Proteins in Cellular Function and in the Biology of Fungi

“…Antigen having the same antigenic epitope in different fungi is potentially recognised by the same antibodies against them, which could inhibit fungal growth and development [ 133 ]. Various Hsps such as Hsp70, Hsp40, Hsp90, and Hsp60 has been shown to be upregulated when different sets of fungi ( C. albicans, P. brasiliensis, and Coccidioides posadasii ) were injected in mice model [ 134 ]. Hsps come under conserved protein domain common in fungi which commonly act as an antigen in fungal infections and showed potential for the development of pan fungal vaccine, providing absence of antigenic region of fungal Hsps in human counterpart.…”

Study of Humoral Responses against Lomentospora/Scedosporium spp. and Aspergillus fumigatus to Identify L. prolificans Antigens of Interest for Diagnosis and Treatment

“…In contrast, sera from mice infected with A. fumigatus reacted with very few proteins of L. prolificans, including enolase. This result is not surprising, since this antigenic protein is highly conserved, showing cross-reactivity between several fungal species [11,46,47].…”