computational methods to inferring cellular networks stat 877 apr 15 th 2014 sushmita roy
TRANSCRIPT
Computational methods to inferring cellular networks
Stat 877Apr 15th 2014Sushmita Roy
Goals for today
• Introduction– Different types of cellular networks
• Methods for network reconstruction from expression– Per-gene vs Per-module methods– Sparse Candidates Bayesian networks – Regression-based methods• GENIE3• L1-DAG learn
• Assessing confidence in network structure
Why networks?
“A system is an entity that maintains its function through the interaction of its parts”– Kohl & Noble
To understand cells as systems: measure, model, predict, refine
Uwe Sauer, Matthias Heinemann, Nicola Zamboni, Science 2007
Different types of networks
• Physical networks – Transcriptional regulatory networks: interactions between
regulatory proteins (transcription factors) and genes– Protein-protein: interactions among proteins– Signaling networks: protein-protein and protein-small molecule
interactions to relay signals from outside the cell to the nucleus
• Functional networks– Metabolic: reactions through which enzymes convert substrates
to products– Genetic: interactions among genes which when perturbed
together produce a significant phenotype than when individually perturbed
Transcriptional regulatory networks
Regulatory network of E. coli.153 TFs (green & light red), 1319 targets
Vargas and Santillan, 2008
A B
Gene C
Transcription factors (TF)
C
A B
• Directed, signed, weighted graph• Nodes: TFs and Target genes• Edges: A regulates B’s expression
level
DNA
Reactions associated with Galactose metabolism
Metabolic networks
MetabolitesN
Ma b
c
O
Enzymes
d
O
M N
KEGG
• Unweighted graph• Nodes: Metabolic enzyme• Edges: Enzymes M and N share a
compound
Protein-protein interaction networks
Barabasi et al. 2003
Yeast protein interaction network
X Y
XY
• Un/weighted graph• Nodes: Proteins• Edges: Protein X physically
interacts with protein Y
Challenges in network biology
Network structure analysis
Network reconstruction/inference (today)
A B
X Y
A B A
X Y
Hubs, degree-distributions,Network motifs
? ? ?Identifying edges and their logic X=f(A,B)
Y=g(B)Node attributes
2
1
Structure Parameters
Network applications f
f
g?
Predicting function and activity of genesfrom network
3
Goals for today
• Introduction– Different types of cellular networks
• Methods for network reconstruction from expression– Per-gene vs Per-module methods– Sparse Candidates Bayesian networks – Regression-based methods• GENIE3• L1-DAG learn
• Assessing confidence in network structure
Computational methods to infer networks
• We will focus on transcriptional regulatory networks– These networks control what genes get activated
when– Precise gene activation or inactivation is crucial for
many biological processes– Microarrays and RNA-seq allows us to systematically
measure gene activity levels • These networks are primarily inferred from gene
expression data
What do we want a model to capture?
X3=ψ(X1,X2)
Function
X1 X2
X3
BOOLEANLINEARDIFF. EQNSPROBABILISTIC…
.
How they determine expression levels?
Sko1
Structure
HSP12
Hot1
Who are the regulators?
Hot1 regulates HSP12
HSP12 is a target of Hot1
Input: Transcription factor level (trans)
HSP12Sko1Hot1
Output: expression levels
Mathematical representations of regulatory networks
X1 X2
X3
f
Output expression of target gene
Models differ in the function that maps regulator input levels to target levels
Input expression/activity of regulators
Rate equations Probability distributions
Boolean Networks Differential equations Probabilistic graphical models
X1 X2
0 0
0 1
1 0
1 1
X3
0
1
1
1
Input OutputX1 X2
X3
Regulatory network inference from expression
Expression-based network inference
Gen
es
Experiments
X2
Structure
X3
X1
X3=f(X1,X2)
Function
X1 X2
X3Expression level of gene i in experiment j
Two classes of expression-based methods
• Per-gene/direct methods (Today)
• Module based methods (Thursday)
X5X3
X1 X2
Module
X3
X1 X2
X5
X3 X4
Per-gene methods
X3
X1 X2
X5
X3 X4
• Key idea: find the regulators that “best explain” expression level of a gene
• Probabilistic graphical methods– Bayesian network
• Sparse Candidates– Dependency networks
• GENIE3, TIGRESS
• Information theoretic methods– Context Likelihood of relatedness– ARACNE
Module-based methods
• Find regulators for an entire module– Assume genes in the same module have the same
regulators• Module Networks (Segal et al. 2005)• Stochastic LeMoNe (Joshi et al. 2008)
Per module
Y2Y1
X1 X2
Module
Goals for today
• Introduction– Different types of cellular networks
• Methods for network reconstruction from expression– Per-gene vs Per-module methods– Sparse Candidates Bayesian networks – Regression-based methods• GENIE3• L1-DAG learn
• Assessing confidence in network structure
Notation
• V: A set of p network components– p genes
• E: Edge set connecting V• G=(V, E). G is the graph we wish to infer• Xv: Random variable, for v ε V
• X={X1,.., Xp}
• D: Dataset of N measurements for X– D: {x1,…xN}
• Θ: Set of parameters associated with the network
Spang and Markowetz, BMC Bioinformatics 2005
Bayesian networks (BN)
• Denoted by B={G, Θ}– G: Graph is directed and acyclic (DAG)– Pa(Xi): Parents of Xi
– Θ: {θ1,.., θp} Parameters for p conditional probability distributions (CPD) P(Xi | Pa(Xi) )
• Vertices of G correspond to random variables X1… Xp
• Edges of G encode directed influences between X1… Xp
A simple Bayesian network of four variables
Adapted from “Introduction to graphical models”, Kevin Murphy, 2001
Random variables:Cloudy ε {T, F}Sprinker ε {T, F}Rain ε {T, F}WetGrass ε {T,F}
A simple Bayesian network of four variables
Conditional probability distributions (CPD)
Adapted from “Introduction to graphical models”, Kevin Murphy, 2001
Random variables:Cloudy ε {T, F}Sprinker ε {T, F}Rain ε {T, F}WetGrass ε {T,F}
Bayesian network representation of a regulatory network
Bayesian network
TARGET (CHILD)
REGULATORS (PARENTS)X1
X2
X3
X1X2
X3P(X3|X1,X2)
Random variables
HSP12Sko1Hot1
Inside the cell
Hot1:
Sko1:
Hsp12:
P(X1) P(X2)
Bayesian networks compactly represent joint distributions
Example Bayesian network of 5 variables
CHILD
PARENTS
X1 X2
X3
X5
X4
No independence assertions
Independence assertions
Assume Xi is binary
Needs 25 measurements
Needs 23 measurements
CPD in Bayesian networks
• The CPD P(Xi|Pa(Xi)) specifies a distribution over values of Xi for each combination of values of Pa(Xi)
• CPD P(Xi|Pa(Xi)) can be parameterized in different ways
• Xi are discrete random variables – Conditional probability table or tree
• Xi are continuous random variables– CPD can be Gaussians or regression trees
• Consider four binary variables X1, X2, X3, X4
Representing CPDs as tables
X1 X2 X3 t f
t t t 0.9 0.1
t t f 0.9 0.1
t f t 0.9 0.1
t f f 0.9 0.1
f t t 0.8 0.2
f t f 0.5 0.5
f f t 0.5 0.5
f f f 0.5 0.5
P( X4 | X1, X2, X3 ) as a tableX4
X1X2
X4
X3
Pa(X4): X1, X2, X3
Estimating CPD table from data
• Assume we observe the following assignments for X1, X2, X3, X4
T F T T
T T F T
T T F T
T F T T
T F T F
T F T F
F F T F
X1 X2 X3 X4
For each joint assignment to X1, X2, X3, estimate the probabilities for each value of X4
For example, consider X1=T, X2=F, X3=T
P(X4=T|X1=T, X2=F, X3=T)=2/4P(X4=F|X1=T, X2=F, X3=T)=2/4
N=7
A tree representation of a CPD
P( X4 | X1, X2, X3 ) as a tree
X1
P(X4 = t) = 0.9
f t
X2
P(X4 = t) = 0.5
f t
X3
P(X4 = t) = 0.5 P(X4 = t) = 0.8
f t
X1X2
X4
X3
Allows more compact representation of CPDs,by ignoring some unlikely relationships.
The learning problems in Bayesian networks
• Parameter learning on known graph structure– Given data D and G, learn Θ
• Structure learning– Given data D, learn G and Θ
Structure learning using score-based search
...
A function of how well B describes the data D
Scores for Bayesian networks
• Maximum likelihood
• Regularized maximum likelihood
• Bayesian score
Decomposability of scores
• The score of a Bayesian network B decomposes over individual variables
• Enables efficient computation of the score change to local changes
Joint assignment to Pa(Xi) in the dth sample
Search space of graphs is huge
• For N variables there are possible graphs
• Set of possible networks grows super exponentially
N Number of networks
3 8
4 64
5 1024
6 32768
Need approximate methods to search the space of networks
Greedy Hill climbing to search Bayesian network space
• Input: D={x1,..,xN}, An initial graph, B0={G0, Θ0}
• Output: Bbest
• Loop until convergence:– {Bi
1, .., Bim} = Neighbors(Bi) by making local changes to Bi
– Bi+1: arg maxj(Score(Bij))
• Termination: – Bbest= Bi
Local changes to Bi
A
B C
D
A
B C
D
add an edge
A
B C
D
delete an edge
Current network
Check for cycles
Bi
Challenges with applying Bayesian network to genome-scale data
• Number of variables, p is in thousands
• Number of samples, N is in hundreds
Extensions to Bayesian networks to handle genome-scale networks
• Sparse candidate algorithm – Friedman, Nachman, Pe’er. 1999
• Bootstrap to identify high scoring graph features– Friedman, Linial, Nachman, Pe’er. 2000
• Module networks (subsequent lecture)– Segal, Pe’er, Regev, Koller, Friedman. 2005
• Add graph priors (subsequent lecture, hopefully)
The Sparse candidate Structure learning in Bayesian networks
• Key idea: Identify k “promising” candidate parents for each Xi
– k<<p, p: number of random variables– Candidates define a “skeleton graph” H
• Restrict graph structure to select parents from H• Early choices in H might exclude other good parents– Resolve using an iterative algorithm
Sparse candidate algorithm
• Input:– A data set D– An initial Bayes net B0
– A parameter k: max number of parents per variable• Output:
– Final B• Loop until convergence
– Restrict• Based on D and Bn-1 select candidate parents Ci
n-1 for Xi
• This defines a skeleton directed network Hn
– Maximize• Find network Bn that maximizes the score Score(Bn;D) among networks satisfying
• Termination: Return Bn
Selecting candidate parents in the Restrict Step
• A good parent for Xi is one with strong statistical dependence with Xi
– Mutual information provides a good measure of statistical dependence I(Xi; Xj)
– Mutual information should be used only as a first approximation• Candidate parents need to be iteratively refined to
avoid missing important dependences
• A good parent for Xi has the highest score improvement when added to Pa(Xi)
Mutual Information
• Measure of statistical dependence between two random variables, Xi and Xj
Mutual information can miss some parents• Consider the following true network
• If I(A;C)>I(A;D)>I(A;B) and we are selecting k<=2 parents, B will never be selected as a parent
• How do we get B as a candidate parent?• If we used mutual information alone to select candidates, we
might be stuck with C and D
A
B C
D
True network
Computational savings in Sparse Candidate
• Ordinary hill climbing– O(2n) possible parent sets– O(n2) initial score change calculations– O(n) for subsequent iterations
• Complexity of learning constrained on a skeleton directed graph – O(2k) possible parent sets– O(nk) initial score change calculations– O(k) for subsequent iterations
Sparse candidate learns good networks faster than hill-climbing
Dataset 1 Dataset 2
100 variables 200 variables
Greedy hill climbing takes much longer to reach a high scoring bayesian network
Scor
e (h
ighe
r is
bett
er)
Some comments about choosing candidates
• How to select k in the sparse candidate algorithm?• Should k be the same for all Xi ?• If the data are Gaussian could be do something better?• Regularized regression approaches can be used to
estimate the structure of an undirected graph• L1-Dag learn provides an alternate
– Schmidt, Niculescu-Mizil, Murphy 2007– Estimate an undirected dependency network Gundir
– Learn a Bayesian network constrained on Gundir
Dependency network
• A type of probabilistic graphical model• As in Bayesian networks has– A graph component– A probability component
• Unlike Bayesian network – Can have cyclic dependencies
Dependency Networks for Inference, Collaborative Filtering and Data visualization Heckerman, Chickering, Meek, Rounthwaite, Kadie 2000
Selecting candidate regulators for the ith gene using regularized linear regression
Xi= X1 …… Xp-1
bi
1
N
1 p-11
N
1
p-1
Regularization term
?? ?…
Xi
Candidates
Everything other than Xi
L1 norm, sparsity imposing Sets many regression coefficients to 0Also called Lasso regression
Learning dependency networks
• Learning: estimate a set of conditional probability distributions, one per variable.
• P(X,|X-j) could be estimated by solving• A set of linear regression problem• Meinhausen & Buhlmann, 2006 • TIGRESS (Haury et al, 2010)
• A set of non-linear regression problems• Non-linearity captured by Regression Tree
(Heckerman et al, 2000)• Non-linearity captured by Random forest• GENIE3, (Huynh-Thu et al, 2010)
Where do different methods rank?
Marbach et al., 2012 Com
mun
ityRa
ndom
Goals for today
• Introduction– Why should we care?– Different types of cellular networks
• Methods for network reconstruction from expression– Per-gene methods
• Sparse Candidates Bayesian networks • Regression-based methods
– GENIE3– L1-DAG learn
– Per-module methods• Assessing confidence in network structure
Assessing confidence in the learned network
• Typically the number of training samples is not sufficient to reliably determine the “right” network
• One can however estimate the confidence of specific features of the network– Graph features f(G)
• Examples of f(G)– An edge between two random variables– Order relations: Is X, Y’s ancestor?
How to assess confidence in graph features?
• What we want is P(f(G)|D), which is
• But it is not feasible to compute this sum
• Instead we will use a “bootstrap” procedure
Bootstrap to assess graph feature confidence
• For i=1 to m– Construct dataset Di by sampling with
replacement N samples from dataset D, where N is the size of the original D
– Learn a network Bi
• For each feature of interest f, calculate confidence
Does the bootstrap confidence represent real relationships?
• Compare the confidence distribution to that obtained from randomized data
• Shuffle the columns of each row (gene) separately.• Repeat the bootstrap procedure
randomize eachrow independently
genes
Experimental conditions
Application of Bayesian network to yeast expression data
• 76 experiments/microarrays• 800 genes• Bootstrap procedure on 200 subsampled
datasets• Sparse candidate as the Bayesian network
learning algorithm
Bootstrap-based confidence differs between real and actual data
f
f
Random
Real
Example of a high confidence sub-network
One learned Bayesian network Bootstrapped confidence Bayesian network
Highlights a subnetwork associated with yeast mating
Summary
• Network inference from expression provides a promising approach to identify cellular networks
• Bayesian networks are one representation of networks that have a probabilistic and graphical component– Network inference naturally translates to learning problems in
Bayesian networks.• Successful application of Bayesian networks to
expression data requires additional considerations– Reduce potential parents
• statistically or using biological knowledge
– Bootstrap based confidence estimation
Linear regression with N inputs
• Y: output•
intercept Parameters/coefficients
Given: Data=
Estimate:
Information theoretic concepts
• Kullback Leibler (KL) Divergence– Distance between two distributions
• Mutual information– Measures statistical dependence between X and Y– Equal to KL Divergence between P(X,Y) and
P(X)P(Y)• Conditional Mutual information– Measures the information between two variables
given a third
KL Divergence
P(X), Q(X) are two distributions over X
Measuring relevance of Y to X
• MDisc(X,Y)– DKL(P(X,Y)||PB(X,Y))
• MShield(X,Y)– I(X;Y|Pa(X))
• Mscore(X,Y)– Score(X;Y,Pa(X),D)
Conditional Mutual Information
• Measures the mutual information between X and Y, given Z
• If Z captures everything about X, knowing Y gives no more information about X.
• Thus the conditional mutual information would be zero.
What do the Bayesian network edges represent?
Spang and Markowetz, BMC Bioinformatics 2005
Is it just correlation? No.
High correlation could be due to any of the three possible regulatory mechanisms