Posttranslational regulation of self-renewal capacity: insights from proteome and phosphoproteome analyses of stem cell leukemia

Abstract

We recently generated 2 phenotypically similar Hoxa9؉Meis1 overexpressing acute myeloid leukemias that differ by their in vivo biologic behavior. The first leukemia, named FLA2, shows a high frequency of leukemia stem cells (LSCs; 1 in 1.4 cells), whereas the second, FLB1, is more typical with a frequency of LSCs in the range of 1 per several hundred cells. To gain insights into possible mechanisms that determine LSC selfrenewal, we profiled and compared the abundance of nuclear and cytoplasmic proteins and phosphoproteins from these leukemias using quantitative proteomics. These analyses revealed differences in proteins associated with stem cell fate, including a hyperactive p38 MAP kinase in FLB1 and a differentially localized Polycomb group protein Ezh2, which is mostly nuclear in FLA2 and predominantly cytoplasmic in FLB1. Together, these newly documented proteomes and phosphoproteomes represent a unique resource with more than 440 differentially expressed proteins and 11 543 unique phosphopeptides, of which 80% are novel and 7% preferentially phosphorylated in the stem cell-enriched leukemia. (Blood. 2012;120(8):e17-e27)

Integrated nuclear proteomics and transcriptomics identifies S100A4 as a therapeutic target in acute myeloid leukemia

“…Although model systems strongly implicate TF dysregulation in this disease, in patient material only a few common transcription factor abnormalities are known (e.g., PML-RARA, RUNX1-ETO, Inv16, CEPBA) and these are mainly associated with particular cytogenetic subsets of AML [36]. Therefore, large scale proteomic technologies are mainly used to quantify changes in protein abundance in in vivo or ES differentiation model systems [37][38][39]. However, the actual transcriptional environment in primary AML patient material characterized by the relative abundance of TF protein expression compared with normal blasts has not yet been described.…”

Proteomic Identification of Novel Secreted Antibacterial Toxins of the Serratia marcescens Type VI Secretion System

“…One milligram of cell lysates were reduced, alkylated, and digested as described above. After sample clean-up using solid phase extraction cartridges (HPE, Waters), phosphopeptides were enriched using TiO 2 chromatography (44–46). Phosphopeptides were analyzed on an Orbitrap Velos Pro, using above LC and mass spectrometry settings.…”

Elucidating Novel Hepatitis C Virus–Host Interactions Using Combined Mass Spectrometry and Functional Genomics Approaches

“…Label-free quantitative proteomics was used to profile protein abundance across sample sets, as reported previously (22,23). Briefly, Mascot peptide identifications were matched to ion intensity (MS peak intensity, minimum threshold: 10,000 counts) extracted from the aligned MS raw data files (tolerances set to m/z ϭ 15 ppm and retention time ϭ 1 min).…”