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Background  Nuclear proteins play critical roles in regulating mRNA transcription and processing, DNA replication, and epigenetic genome modification. Therefore, the ability to monitor changes in nuclear proteins is helpful not only to identify important regulatory proteins but also to study the mechanisms of actions of nuclear proteins. However, no effective methods have been developed yet. Rye is strongly resistant to various biotic and abiotic stresses; however, few genes have been functionally characterized to date due to the complexity of its genome and a lack of genomic sequence information.    Results  We developed an integrative Nuclear Transportation Trap (iNTT) system that includes an improved nuclear transportation trap and utilizes the “after suppression subtraction” method. Oligonucleotides encoding a nuclear localization signal (NLS) or a transcription factor, GmAREB, were inserted into pLexAD or pLexAD-NES, respectively, and then transformed into yeast cells (EGY48). We showed that the pLexAD vector expressing a cDNA library in the iNTT system was more efficient for screening than the vector pLexAD-NES, which has previously been used in an NTT system. We used the iNTT system to screen a cDNA library of cold-treated rye. A total of 241 unique genes were identified, including 169 differentially expressed proteins; of these, 106 were of known and 63 were of unknown function. Moreover, 82 genes (49 %) among the 169 differentially expressed genes were predicted to contain an NLS domain. Thirty-three (31 %) of the 106 functionally known proteins have DNA-binding activity. To test the specificity of the nuclear proteins identified using the iNTT screen, four of the proteins differentially expressed in response to temperature stress, ScT1 (a heat shock protein), ScT36 (a MYB-like transcription factor), ScT133 (an ERF-like transcription factor) and ScT196 (a protein of unknown function), were studied in more depth. These proteins were shown to exclusively localize to the nucleus, and their expression levels were increased in response to low-temperature stress. To identify the function of these screened nuclear proteins,  ScT1-  and  ScT36- transgenic  Arabidopsis  plants were constructed, and  ScT1  or  ScT36  overexpression was found to enhance tolerance to high-temperature or freezing stresses, respectively.    Conclusions  The newly developed iNTT system provides an effective method for identifying nuclear-targeted proteins and monitoring induced expression levels.  ScT1  and  ScT36  might be good candidate genes for improving the stress tolerance of plants by genetic transformation.    Electronic supplementary material  The online version of this article (doi:10.1186/s12864-016-2548-y) contains supplementary material, which is available to authorized users.

bmc genomics

An iNTT system for the large-scale screening of differentially expressed, nuclear-targeted proteins: cold-treatment-induced nucleoproteins in Rye (Secale cereale L.)