An Approach to Function Annotation for Proteins of Unknown Function (PUFs) in the Transcriptome of Indian Mulberry

The modern sequencing technologies are generating large volumes of information at the transcriptome and genome level. Translation of this information into a biological meaning is far behind the race due to which a significant portion of proteins discovered remain as proteins of unknown function (PUFs). Attempts to uncover the functional significance of PUFs are limited due to lack of easy and high throughput functional annotation tools. Here, we report an approach to assign putative functions to PUFs, identified in the transcriptome of mulberry, a perennial tree commonly cultivated as host of silkworm. We utilized the mulberry PUFs generated from leaf tissues exposed to drought stress at whole plant level. A sequence and structure based computational analysis predicted the probable function of the PUFs. For rapid and easy annotation of PUFs, we developed an automated pipeline by integrating diverse bioinformatics tools, designated as PUFs Annotation Server (PUFAS), which also provides a web service API (Application Programming Interface) for a large-scale analysis up to a genome. The expression analysis of three selected PUFs annotated by the pipeline revealed abiotic stress responsiveness of the genes, and hence their potential role in stress acclimation pathways. The automated pipeline developed here could be extended to assign functions to PUFs from any organism in general. PUFAS web server is available at http://caps.ncbs.res.in/pufas/ and the web service is accessible at http://capservices.ncbs.res.in/help/pufas.现代测序技术正从转录组与基因组层面产生海量信息。然而将此类信息转化为生物学意义的进程远远滞后，致使大量新发现的蛋白质仍属于功能未知蛋白（proteins of unknown function, PUFs。当前针对PUFs功能意义的探索受限于缺乏简便易用的高通量功能注释工具。本文报道了一种为功能未知蛋白推定功能的方法，针对的是作为家蚕宿主的多年生乔木——桑树转录组中鉴定出的PUFs。本研究使用了全植株水平下暴露于干旱胁迫的叶片组织所产生的桑树PUFs，通过基于序列与结构的计算分析预测了这些PUFs的可能功能。为实现PUFs的快速简便注释，我们开发了整合多种生物信息学工具的自动化流程，命名为PUFs注释服务器（PUFs Annotation Server, PUFAS），该服务器还提供了适用于基因组级大规模分析的Web服务应用程序编程接口（Application Programming Interface, API）。对通过该流程注释的三个选定PUFs进行表达分析后发现，这些基因响应非生物胁迫，因此它们可能在胁迫适应通路中发挥潜在作用。本文开发的自动化流程原则上可推广应用于为任意生物的PUFs分配功能。PUFAS网页服务器的访问地址为http://caps.ncbs.res.in/pufas/，其Web服务的访问地址为http://capservices.ncbs.res.in/help/pufas。