Study of the Mtf A Transcription Factor in the Opportunistic Fungal Pathogen Aspergillus fumigatus

Ref ID: 19512

Author:

TD Smith1*, AM Calvo1

Author address:

1Biology, Northern Illinois University, DeKalb, United States of America

Full conference title:

6th Advances Against Aspergillosis 2014

Abstract:

Purpose:
Aspergillus fumigatus is the leading causative agent of invasive aspergillosis. The number of cases
are on the rise with mortality rates being as high as 90%. This study covers a novel regulatory gene
in A. fumigatus that could potentially be used to design novel anti-fungal therapies. In this work we
investigated the role of the transcription factor gene mtfA in A. fumigatus. Our recent study of the mtfA
ortholog in the model fungus Aspergillus nidulans revealed that mtfA is a master genetic regulator that
governs sexual and asexual development as well as the production of various secondary metabolites. In
the present study we show that in A. fumigatus mtfA controls veA and laeA expression and plays a role in
growth, development gliotoxin production, protease activity and virulence.
Methods:
In order to characterize mtfA in A. fumigatus deletion mtfA, complementation, and over-expression strains
were generated. Colony growth was determined by point inoculating the strains on Czapek-Dox medium
and allowing them to grow for five days. Conidiation data was collected from top-agar inoculated cultures
at 48 and 72 hrs. For all gene expression data, strains were inoculated in liquid stationary cultures and
samples collected at 48 h and 72 h. Gene expression was measured by qRTPCR. Gliotoxin production
was analyzed on culture filtrates at 3 d and 5 d by HPLC. Protease activity was measured using an Azo-
Casein Assay. Virulence studies were done using the Galleria mellonella infection model.
Results:
Deletion and over-expression of mtfA in A. fumigatus resulted in a decrease in colony growth (deletion
of mtfA resulted in a 18% decrease in colony diameter while over expression resulted in a 24%
decrease). Complementation of the mtfA deletion strain with the mtfA wilt-type allele was able to partially
restore wild-type colony growth. Along with a decrease in colony growth, both deletion and overexpression
of mtfA resulted in a decrease in conidiation. Our studies also showed that mtfA is necessary
for proper levels of veA and of laeA expression. laeA expression was most affected (over-expression of
mtfA in A. fumigatus resulted in a two fold increase in laeA expression levels at both 48 h and 72 h after
inoculation). Additionally, protease activity levels in the deletion mtfA mutant were decreased 50% with
respect to the wild-type strain, while mtfA over-expression resulted in an increase in protease activity up
to 116%. With respect to gliotoxin, both deletion and over-expression of mtfA resulted in an increase in
the production of this compound at 72 h and 120 h. At 72 h the mtfA deletion strain showed a 5.7 fold
increase in gliotoxin with respect to wild-type and over-expression showed a 1.8 fold increase. At 120
h deletion mtfA showed a 6.2 fold increase in gliotoxin production and over-expression levels were 8.7
fold greater. Although an increase in gliotoxin was observed, in the Galleria infection model deletion of
mtfA resulted in a decrease in virulence. Thirty percent of the larvae infected with the deletion mutant
were still alive at the end of the study whereas only 6% of those infected with the isogenic wild-type
strain remained living. Over-expression showed no statistical difference in survival rate with respect to
wild type in this study.
Conclusion:
Our studies have revealed that mtfA plays a role in growth and conidiation in the opportunistic pathogen
Aspergillus fumigatus. Interestingly, mtfA regulates the expression levels of the global regulators veA and
laeA, known to control development and secondary metabolism in many fungal species. Proper levels of
mtfA are required for wild-type levels of protease activity. Gliotoxin production in A. fumigatus is also
affected by mtA. Importantly, deletion of mtfA resulted in decreased virulence in a Galleria infection
model.
NOTE: THIS ABSTRACT AS BEEN SELECTED FOR ORAL PRESENTATION.

Abstract Number: 40

Conference Year: 2014

Link to conference website: http://www.AAA2014.org

New link: NULL

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