| 1. |
|
|
| 2. |
- Mahmoudi, Salah, et al.
(författare)
-
WRAP53 Is Essential for Cajal Body Formation and for Targeting the Survival of Motor Neuron Complex to Cajal Bodies
- 2010
-
Ingår i: PLoS biology. - : Public Library of Science (PLoS). - 1544-9173 .- 1545-7885. ; 8:11, s. e1000521-
-
Tidskriftsartikel (refereegranskat)abstract
- The WRAP53 gene gives rise to a p53 antisense transcript that regulates p53. This gene also encodes a protein that directs small Cajal body-specific RNAs to Cajal bodies. Cajal bodies are nuclear organelles involved in diverse functions such as processing ribonucleoproteins important for splicing. Here we identify the WRAP53 protein as an essential factor for Cajal body maintenance and for directing the survival of motor neuron (SMN) complex to Cajal bodies. By RNA interference and immunofluorescence we show that Cajal bodies collapse without WRAP53 and that new Cajal bodies cannot be formed. By immunoprecipitation we find that WRAP53 associates with the Cajal body marker coilin, the splicing regulatory protein SMN, and the nuclear import receptor importin beta, and that WRAP53 is essential for complex formation between SMN-coilin and SMN-importin beta. Furthermore, depletion of WRAP53 leads to accumulation of SMN in the cytoplasm and prevents the SMN complex from reaching Cajal bodies. Thus, WRAP53 mediates the interaction between SMN and associated proteins, which is important for nuclear targeting of SMN and the subsequent localization of the SMN complex to Cajal bodies. Moreover, we detect reduced WRAP53-SMN binding in patients with spinal muscular atrophy, which is the leading genetic cause of infant mortality worldwide, caused by mutations in SMN1. This suggests that loss of WRAP53-mediated SMN trafficking contributes to spinal muscular atrophy.
|
|
| 3. |
- Weibrecht, Irene, et al.
(författare)
-
Proximity ligation assays : a recent addition to the proteomics toolbox
- 2010
-
Ingår i: Expert Reviews of Proteomics. - 1478-9450. ; 7:3, s. 401-409
-
Forskningsöversikt (refereegranskat)abstract
- An essential skill for every researcher is to learn how to select and apply the most appropriate methods for the questions they are trying to answer. With the extensive variety of methods available, it is increasingly important to scrutinize the advantages and disadvantages of these techniques prior to making a decision on which to use. In this article, we describe an approach to evaluate methods by reducing them into subcomponents. This is exemplified by a brief description of some commonly used proteomics methods. The same approach can also be used in method development by rearranging subcomponents in order to create new methods, as demonstrated with the development of proximity ligation assays (PLAs). PLA is a method as designed in our laboratory for detection of proteins, protein-protein interactions and post-translational modifications. Fundamentally, protein-recognition events are converted into detectable DNA molecules. The technique uses protein DNA conjugates as binders for the targets of interest. Binding of two or more conjugates to the target results in assembly of an assay-specific DNA molecule. Subsequent amplification of the DNA molecule generates a signal that can be detected using PCR, for detection of minute amounts of proteins in serum, or standard fluorescence microscopy for detection of protein protein interactions in tissue sections. Lastly, we apply the approach of recombining subcomponents to develop a few novel hypothetical methods hoping this might stimulate the readers to utilize this approach themselves.
|
|
