Active and Repressive Chromatin-Associated Proteome after MPA Treatment and the Role of Midkine in Epithelial Monolayer Permeability.
Mycophenolic acid (MPA) is prescribed to maintain allografts in organ-transplanted patients. However, gastrointestinal (GI) complications, particularly diarrhea, are frequently observed as a side effect following MPA therapy. We recently reported that MPA altered the tight junction (TJ)-mediated barrier function in a Caco-2 cell monolayer model system. This study investigates whether MPA induces epigenetic changes which lead to GI complications, especially diarrhea.


METHODS
We employed a Chromatin Immunoprecipitation-O-Proteomics (ChIP-O-Proteomics) approach to identify proteins associated with active (H3K4me3) as well as repressive (H3K27me3) chromatin histone modifications in MPA-treated cells, and further characterized the role of midkine, a H3K4me3-associated protein, in the context of epithelial monolayer permeability.


RESULTS
We identified a total of 333 and 306 proteins associated with active and repressive histone modification marks, respectively. Among them, 241 proteins were common both in active and repressive chromatin, 92 proteins were associated exclusively with the active histone modification mark, while 65 proteins remained specific to repressive chromatin. Our results show that 45 proteins which bind to the active and seven proteins which bind to the repressive chromatin region exhibited significantly altered abundance in MPA-treated cells as compared to DMSO control cells. A number of novel proteins whose function is not known in bowel barrier regulation were among the identified proteins, including midkine. Our functional integrity assays on the Caco-2 cell monolayer showed that the inhibition of midkine expression prior to MPA treatment could completely block the MPA-mediated increase in barrier permeability.


CONCLUSIONS
The ChIP-O-Proteomics approach delivered a number of novel proteins with potential implications in MPA toxicity. Consequently, it can be proposed that midkine inhibition could be a potent therapeutic approach to prevent the MPA-mediated increase in TJ permeability and leak flux diarrhea in organ transplant patients.
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany. niamat.khattak@gmail.com.|Department of Biotechnology &amp; Genetic Engineering, Kohat University of Science and Technology, Kohat 26000, Khyber Pakhtunkhwa, Pakistan. niamat.khattak@gmail.com.
Niamat
N
Khan
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany. christof.lenz@mpibpc.mpg.de.|Bioanalytical Mass Spectrometry, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany. christof.lenz@mpibpc.mpg.de.
Christof
C
Lenz
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany. lutz.binder@med.uni-goettingen.de.
Lutz
L
Binder
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany. krishna.if1@gmail.com.
Dasaradha Venkata Krishna
DV
Pantakani
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany. asif@med.uni-goettingen.de.
Abdul R
AR
Asif
D018938:Caco-2 Cells; D002843:Chromatin / Q000378:metabolism; D042002:Chromatin Assembly and Disassembly; D047369:Chromatin Immunoprecipitation / Q000379:methods; D044127:Epigenesis, Genetic; D006657:Histones / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000187:drug effects* / Q000378:metabolism; D008745:Methylation; D000078792:Midkine; D009173:Mycophenolic Acid / Q000494:pharmacology*; D009414:Nerve Growth Factors; D016377:Organ Transplantation / Q000009:adverse effects*; D010539:Permeability / Q000187:drug effects*; D020543:Proteome; D019108:Tight Junctions / Q000187:drug effects*
D002843:Chromatin; D006657:Histones; C066254:MDK protein, human; D009414:Nerve Growth Factors; D020543:Proteome; D000078792:Midkine; D009173:Mycophenolic Acid
iMDK; midkine; mycophenolic acid; tight junctions