A Critical Appraisal of Carlisle et al

Siti Nor Atikah Mohamad
BMS3007 Critical Appraisal
A Critical Appraisal of Carlisle et al (2009), ‘Expression Levels
of a Filament-specific Transcriptional Regulator are
Sufficient to Determine Candida albicans Morphology and
Carlisle et al conducted a study on a recently identified filament-specific transcriptional regulator of
Candida albicans hyphal extension and virulence, UME6 to address relative contribution of C.albicans
morphologies to virulence, which for many years has been inadequately known. This is of clinical interest
and importance; C.albicans as the most common human fungal pathogen is responsible for wide-variety
of mucosal and systemic infection worldwide and is 4th leading cause of hospital-acquired bloodstream
infection in USA (1). The study is relevant and significant to date, provided the evidence that typical
C.albicans infection contains a mixture of yeast, pseudohyphal and hyphal filaments (2). In addition,
earlier findings have shown that C.albicans hyphal form involved in various virulence-related process
during infection in the host including ‘the invasion of epithelial layers, thigmotropism, and lysis of
macrophages and neutrophils’ (3, 4). Therefore the study aims to investigate the regulatory mechanism
underlying hyphae and pseudohyphal transformation in C.albicans and the role of hyphae formation in
C.albicans virulence. To address this, the study generated C.albicans strain that can be genetically
manipulated to grow in hyphal form in the absence of filament-inducing conditions and this was achieved
by inducement of constitutive high-level of UME6 expressions (5) . The result strongly suggest that the
morphology shifting to hyphal form and/or increasing hyphal-specific gene expression is sufficient to
improve virulence potential during course of infection (5).
The introduction gives extensive background information on C.albicans infections, including the fact that
immunocompromised individuals are particularly susceptible to infection (5). This highlighted the
significance of this study to provide a powerful tool to address the regulatory mechanism involved to
promote virulence. The paper explains the different morphologies of the C.albicans that are reversible
from a single yeast cell, to psuedohyphal and hyphal filaments and emphasizes the extensive phenotypic
differences between hyphal and psuedohyphal filament cells. The paper therefore recognizes the
importance of identifying the genetic determinants for each growth of these filaments and the specific
role that each plays to cause virulence as less is known. Briefly, the introduction gives apparent
information of what the group want from the study and the aim is put forward well.
In the material and method section, the preparation of strains and DNA construction is presented quiet
thoroughly containing the important steps involved such as the integration event to generate the tetOUME6 strain. The study also takes necessary step to confirm the integration event showing the reader
that attention is taken during the preparation.
Siti Nor Atikah Mohamad
BMS3007 Critical Appraisal
For virulence experiments and histology, the method used to demonstrate virulence in mouse model of
systemic candidiasis is simple and effective. 3 days prior to infection, 6-8 old weeks of BALB/c mice were
placed on drinking water with removal or addition of dox. The difference in virulence between +DOX and
–DOX groups is proven to be statistically significant by carrying out Kaplan-Meier test, taken into account
that the sample size used is very small. The histology experiment on the kidney of the deceased mouse to
visualize the morphologies of the fungal cell is clearly a necessary step to confirm the expressions levels
of UME6 alone are sufficient to determine C.albicans morphology. However, the paper has not mentioned
the ethical approval was granted.
Perhaps the most essential part of all of the experiments in the study is the using of tetracyclineregulatable gene expression system that was successfully used by several previous studies. This allow the
study to induce constitutive high-level of UME6 expression and enable to demonstrate the effect on the
C.albicans morphology and virulence (5). By using this system, the gene expression during infection in
mouse model is controllable with addition or removal of Dox (5).
The data of the results was expressed coherently with each results is presented with corresponding figure
and diagram. For example the paper include the histology visualization of the kidney tissues for all of the
samples which allows for comparison.
Perhaps the paper most significant findings was the evidence that expression levels of UME6 are sufficient
to determine the growth of both pseudohyphal and hyphal morphologies of C.albicans. This challenge the
current paradigm that genetically distinct developmental states are specify for each. Hence, the paper
suggests a paradigm shift in the field that the “transitions between C.albicans morphologies is controlled
by a common transcriptional regulatory mechanism in a dosage-dependent manner” (5). The result
proposed a model in which the expression level of UME6 determine the morphologies transition by
“controlling the expression levels of overlapping subsets of genes in the C.albicans filamentous growth
program” (5). At low level of UME6 expression, a fraction of the filament-specific transcripts (ECE1, PHR1,
and HCG1) are induced, specifying growth in the pseudohyphal form. While the increasing levels of
expression cause increase inducement of the same set of genes resulting in the growth of hyphal form.
The study should also be credited for being the first team to carry out inoculations using yeast form cells
which play an important role in dissemination. Therefore, it is reasonable to believe that the experiment
is more closely equivalent to an actual infection.
Despite these significant findings, one should not always generalized findings across animal models
although the mouse physiology is the closest to humans. The study can be greatly improved if the sample
size is bigger as a total of 8 mouse could be said as insufficient.
Although the relationship between pseudohyphal and hyphal morphologies still remain elusive, the study
has reveal pseudohyphal form as an intermediate morphology which is unique to C.albicans. Considering
C. albicans feature as one of the most highly evolved fungal pathogen, this propose that the intermediate
Siti Nor Atikah Mohamad
BMS3007 Critical Appraisal
morphology may benefited evolutionary advantage during infection (5). Further studies ought to be
undertaken to provide more comprehensive understanding of these regulatory mechanism as mentioned
in the paper. Interestingly, a study has revealed that UME6 expression enhances biofilm formation via
Hgc1 and Sun41-dependent mechanism, by provided a strong correlation between increase hyphal
growth and enhanced biofilm formation (6). In conclusion, the study makes large step in defining the role
of hyphal form to virulence as well as the regulatory mechanism involved even though not entirely
conclusive. The study has led pathway to a more clinically significant studies and perhaps more in the
future to provide better treatment for C.albicans infection.
Words Count: 1030
1. Edmond MB, et al. (1999) Nosocomial bloodstream infections in United States hospitals:A threeyear analysis. Clin Infect Dis 29:239–244
2. Odds FC (1988) Candida and Candidosis (Baillie` re Tindall, London).
3. Lo HJ, et al. (1997) Nonfilamentous C. albicans mutants are avirulent. Cell 90:939–949.
4. Kumamoto CA, Vinces MD (2005) Contributions of hyphae and hypha-co-regulated genes to
Candida albicans virulence. Cell Microbiol 7:1546–1554.
5. Carlisle PL, et al (2009) Expression Levels of a Filament-specific Transcriptional Regulator are
Sufficient to Determine Candida albicans Morphology and Virulence. Proceedings of the National
Academy of Sciences(PNAS) 2:599-604
6. Banerjee et al (2013) Expression of UME6, A Key Regulator of Candida albicans Hyphal
Development, Enhances Biofilm Formation via Hgc1- and Sun41-Dependent Mechanisms.
Eukaryotic Cell 2:224-232
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