Genetic analysis of the Candida albicans biofilm transcription factor network using simple and complex haploinsufficiency

Biofilm formation by Candida albicans is a key aspect of its pathobiology and is regulated by an integrated network of transcription factors (Bcr1, Brg1, Efg1, Ndt80, Rob1, and Tec1). To understand the details of how the transcription factors function together to regulate biofilm formation, we used a systematic genetic interaction approach based on generating all possible double heterozygous mutants of the network genes and quantitatively analyzing the genetic interactions between them. Overall, the network is highly susceptible to genetic perturbation with the six network heterozygous mutants all showing alterations in biofilm formation (haploinsufficiency). In addition, many double heterozygous mutants are as severely affected as homozygous deletions. As a result, the network shows properties of a highly interdependent ‘small-world’ network that is highly efficient but not robust. In addition, these genetic interaction data indicate that TEC1 represents a network component whose expression is highly sensitive to small perturbations in the function of other networks TFs. We have also found that expression of ROB1 is dependent on both auto-regulation and cooperative interactions with other network TFs. Finally, the heterozygous NDT80 deletion mutant is hyperfilamentous under both biofilm and hyphae-inducing conditions in a TEC1-dependent manner. Taken together, genetic interaction analysis of this network has provided new insights into the functions of individual TFs as well as into the role of the overall network topology in its function.白色念珠菌（Candida albicans）的生物膜形成是其致病生物学的关键特征，受由转录因子（transcription factors）Bcr1、Brg1、Efg1、Ndt80、Rob1及Tec1组成的整合调控网络所调控。为阐明这些转录因子协同调控生物膜形成的具体机制，本研究采用了系统性遗传互作分析方法：通过构建该调控网络基因的全部可能双杂合突变体，并对其间的遗传互作进行定量分析。整体而言，该调控网络对遗传扰动具有高度敏感性：6种网络基因杂合突变体均表现出生物膜形成异常（单倍体不足，haploinsufficiency）。此外，诸多双杂合突变体的表型严重程度与纯合缺失突变体相当。由此可见，该调控网络呈现出高度互依的‘小世界网络（small-world network）’特性，其运行效率极高但缺乏鲁棒性。此外，上述遗传互作数据表明，TEC1是该调控网络的核心组分，其表达对其他网络转录因子功能的微小扰动极为敏感。本研究同时发现，ROB1的表达既依赖于自身调控，也依赖于与其他网络转录因子的协同互作。最后，NDT80杂合缺失突变体在生物膜形成及菌丝诱导条件下均呈现菌丝过度形成的表型，且该表型依赖于TEC1的调控。综上，对该调控网络的遗传互作分析，不仅为单个转录因子的功能提供了新见解，也揭示了整体网络拓扑结构在其调控功能中的作用。