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Abstract: Integration of exogenous sequence into an intact genome may cause some artificial phenotype or unspecific observations. We noticed that there is unspecific vascular expression when using fli1a:EGFP transgenic embryos for whole-mount in situ hybridization (WISH) experiments. We therefore tested whether the residual vector sequence contained in the fli1a:EGFP transgene or the integration of transgene into the genome may cause this expression ‘noise’ and/or deregulation of gene expression at a genome-wide level. RNA probes were synthesized using two different methods, i.e. vector-based and PCR-based. The vector-based dnmt3 probe showed unspecific vascular expression in fli1a:EGFP embryos, but not in wildtype embryos, by WISH. Moreover, we also found that compared to that in wildtype, there were alterations in gene expression at whole-genome level in the fli1a:EGFP embryos. Our finding that the vector sequence contained in the fli1a:EGFP genome causes unspecific vascular expression by WISH and the genome-wide expression profiling is altered in fli1a:EGFP embryos strongly argue that extra caution should be taken for data interpretation when using transgenics, such as fli1a:EGFP, in developmental biology studies. [Copyright &y& Elsevier]

Abscisic acid (ABA) is an important plant hormone that mediates abiotic stresses in plant growth and development. A number of E3 ligases have been reported to be involved in ABA signaling pathway. In this study, we identified a C3H2C3 RING-type E3 ligase, Arabidopsis thaliana Tόxicos en Levadura 61 (ATL61), which regulated drought stress in planta. Enzyme assay in vitro demonstrated that ATL61 had E3 ubiquitin ligase activity, while point mutation of ATL61H109A, H122A (mATL61) abolished its E3 ubiquitin ligase activity. ATL61 overexpression plants exhibited ABA hypersensitivity and were more tolerant to drought, while the atl61 mutant plants were insensitive to ABA. Moreover, mATL61 overexpression lines exhibited similar ABA-related phenotypes with wild type (WT) plants. The transcript abundances of ABA-mediated drought stress-related genes RD20 and RD22 were higher in ATL61 overexpression plants than those in WT, atl61, and mATL61 plants. Our results indicated that ATL61 acted as a positive regulator in the ABA-mediated drought stress response. • ATL61 is a C3H2C3 RING type E3 ligase with necessary H109 and H122 residues. • ATL61 is induced by ABA and other abiotic stresses and involved in ABA-mediated plants tolerance to drought. • E3 ligase activity of ATL61 was critical to the function of ATL61 in ABA signaling pathway. [ABSTRACT FROM AUTHOR]

The calcium-dependent protein kinases (CDPKs) play vital roles in plant response to various environmental stimuli. Here, we investigated the function of Arabidopsis AtCPK1 in response to salt and drought stress. The loss-of-function cpk1 mutant displayed hypersensitive to salt and drought stress, whereas overexpressing AtCPK1 in Arabidopsis plants significantly enhanced the resistance to salt or drought stress. The reduced or elevated tolerance of cpk1 mutant and AtCPK1 -overexpressing lines was confirmed by the changes of proline, malondialdehyde (MDA) and H 2 O 2 . Real-time PCR analysis revealed that the expression of several stress-inducible genes ( RD29A, COR15A, ZAT10, APX2 ) down-regulated in cpk1 mutant and up-regulated in AtCPK1 -overexpressing plants. These results are likely to indicate that AtCPK1 positively regulates salt and drought stress in Arabidopsis. [ABSTRACT FROM AUTHOR]