Abstract
The immune system is unique in its dynamic interplay between numerous cell
types. However, a system-wide view of how immune cells communicate to
protect against disease has not been established. Here, we applied
high-resolution mass spectrometry-based proteomics to characterize 28
primary human hematopoietic cell populations in steady and activated states
at a depth of > 10,000 proteins in total. Protein copy numbers reveal a
specialization of immune cells for ligand and receptor expression, thereby
connecting distinct immune functions. By integrating total and secreted
proteomes, we establish a ‘social’ network of communicating
cells and discover fundamental structures of intercellular information
exchange as well as novel connections between cell types. Our publicly
accessible proteomic resource provides a framework for the orchestration of
cellular interplay and a reference for altered communication associated with
pathology. For more details click
here
Protein Copy Numbers
Upper left panel:
Protein rank plot. Proteins are ranked by copy number with the selected
protein highlighted in blue.
Upper right panel: Copy
number profile plot. Horizontal line indicates mean copy number. Error bars
are +/- SD.
Lower left panel: Module
expression profile of selected protein. Colors represent correlation to
module eigengene. Selected protein is displayed in black.
Lower right panel: Module cell-type piechart. Colors represent corresponding cell type and
area relative expression.
For more details see ‘Help Me’.
Communication Structures
Communication wheel with cell types (outer circle); receptor and ligand
gene names (middle circle) and their expression levels (inner circle).
Connecting lines indicate intercellular connections. Connection color code:
Ligand-receptor connections above the threshold are colored by the cell type
color of the cell type expressing the ligand; connection below the threshold
are in grey; receptor-receptor connections are in black.
For more details see ‘Help Me’.
Pairwise cell-type comparisons
Pairwise comparisons of complete proteomes visualized as Volcano plots. T-
test differences are plotted as fold-change (x-axis, in log2) and p-value
(y-axis, in -log10). Proteins with significantly different expression levels
(two-tailed Welch’s t-test, S0=1, FDR<5%) are indicated in red. The
false discovery rate (FDR) can be adjusted. Selected proteins are named and
visualized by blue dot.
For more details see ‘Help Me’.
Protein Copy Numbers
On this page you can visualize individual protein abundances. Select your
favorite protein from the drop-down menu or search for the gene name in the
textbox (‘look up protein’). On the page you see four panels.
The
first panel displays the median copy number of all proteins (grey) derived
from all different cell types in log10 scale. The selected protein is
highlighted by a blue dot. In case the protein was assigned to a module, the
module number is displayed below the gene name. The second panel shows the
mean copy number of the selected protein across all measured cell types.
Error bars are +/- sd (n=4, or n=3, see Online Methods for more details). In
the third panel the module profile is shown. The black line represents the
profile of the selected protein, all other proteins are colored according to
their distance to the selected profile eigengene (blue to red, low to high
Pearson correlation coefficient). In the fourth panel, the module profile is
displayed as a pie. The slice sizes scale according to the module eigengene
values and are colored according to the cell type color code on the
left.
Communication Structures
Menu 1: Plot
On this page you can visualize communication structures as circos plots using the ‘circlize’ R package. Select your favorite protein from the drop-down menu or search for the gene name in the textbox. After your selection, the left panel is updated with potential connection partners, ranked by the STRINGdatabase score in decreasing order. By default, the protein with the highest STRING score is preselected, but you can of course select a different interacting protein to be visualized. If you want to add another protein, use the tick-box ‘add another protein?’, For timely visualization we recommend to not choose too many proteins at the same time. After selecting all proteins press ‘update wheel’ to display the communication wheel. You can also modify the connection threshold (‘cut-off’) to visualize only a subset of all possible connection. The threshold ranges from 0 to 1 (0 will display all connections and 1 no connections; 0.47 was used in the publication and is set as default). Ligand-receptor connections above the threshold are colored by the cell type color of the cell type expressing the ligand; connection below the threshold are indicated in grey; receptor-receptor connections in black. You can download the communication wheel plot in pdf format (‘download plot’).
Menu 2: Table
On this page, the information of the communication wheel is stored, which you can download (‘download table’ bottom, top left). The table contains all edge information of the selected proteins and consists of 6 columns (CellType, Gene.names, rank, experiment score, Gene.names_2, CellType_2). Rank: connection score (see Online Methods for detailed description); experiment score: STRINGdatabase score (experimental evidence only). You can search the table for specific connections (top), filter individual columns for specific terms (bottom), and specify or sort the number of entries and columns.
Pairwise Comparisons
Menu 1: Plot
On this page you can explore and visualize proteome differences by volcano
plots (x-axis: fold change [log2], y-axis: p-value [-log10]). Select your
two favorite cell types from the drop-down menu or search for their names in
the textbox. (‘first cell type’ and ‘second cell
type’). You can adjust the false discovery rate (FDR) (‘FDR
cutt-off’, the default setting of 0.05 is a good starting point) for
pairwise proteome comparisons. By ticking ‘show outlier names’
all proteins below the FDR cut-off are shown with their gene name - by
default this option is disabled. For a timely use of this option, we
recommend settings with not too many significant proteins (red dots). You
can further highlight individual proteins (‘indicate protein’)
in the plot with blue dots, but not more than two at a time (‘add
another protein?’). After setting the parameters, update the volcano
plot (‘update volcano’).
Manual selection tool:
You can select proteins directly inside the plot: Just draw a rectangle
inside the volcano plot with the mouse cursor and move it around. Points
inside the rectangle are listed in the table (‘manual
selection’) below the volcano plot. The table contains 5 columns:
Gene.names, Majority.protein.IDs (Uniprot IDs), p.value, q.value, diff
(fold change).
Menu 2: Table
The information of all proteins in the displayed volcano plot is stored in this table, which you can download (‘download table’, topleft). The table contains 5 columns: Gene.names, Majority.protein.IDs (Uniprot IDs), p.value, q.value, diff (foldchange). You can search the table for specific connections (top), filter individual columns for specific terms (bottom) and specify or sort the number of entries and columns.
Suggestions, questions or just want to chat with us?
For more analyses and details on the methods, please read our paper
Daniel Hornburg 'Dansen'
hornburg [at] biochem [dot] mpg [dot] de
hornburg
[at] stanford [dot] edu