luck et al

What is the main goal of the HuRI project described in this paper?

To generate a systematic, comprehensive reference map of the human binary protein-protein interactome (HuRI) to study cellular organization, genome function, and genotype-phenotype relationships

How many protein-protein interactions (PPIs) and proteins does HuRI contain?

Approximately 53,000 PPIs involving 8,275 proteins

How does the size of HuRI compare to previous high-quality curated interactions from small-scale studies (Lit-BM)?

HuRI has about four times as many PPIs as Lit-BM

What is the name of the human ORFeome version used in this study, and how many protein-coding genes does it cover?

hORFeome v9.1 covers 17,408 protein-coding genes (~90% of the protein-coding genome)

Which experimental assay was primarily used for systematic screening of binary PPIs?

Yeast two-hybrid (Y2H) assay

How many different Y2H assay versions were used, and why?

Three versions; they are complementary, increasing sensitivity and detecting more PPIs than using a single version

What fraction of direct contact pairs within protein complexes does HuRI capture compared to Lit-BM?

90% for HuRI versus 81% for Lit-BM (P = 0.019, two-sided Fisher's exact test)

What is the estimated coverage of HuRI relative to the entire human binary protein interactome?

Approximately 2-11% of the total binary protein interactome

What is a profile similarity network (PSN), and what does it reveal?

A network where proteins with similar interaction partners are linked; it reveals functional relationships between proteins even when sequence identity is low

Give an example of a protein pair with low sequence identity but high interaction profile similarity mentioned in the paper.

TMEM258 and C19ORF18 (10% sequence identity but share 80% of their interaction partners)

How does HuRI coverage compare to Lit-BM for genes with few publications?

HuRI covers the genome-by-genome space more uniformly, while Lit-BM is highly biased toward the most studied genes

What percentage of HuRI PPIs involving the best-studied genes are already in Lit-BM?

Approximately 40%

What confounding factors correlate with protein interaction degree in literature-curated networks but not in HuRI?

Protein popularity (publication count) and endogenous expression level

After correcting for popularity and expression level, what happens to the correlation between interaction degree and gene essentiality?

The correlation is substantially reduced, suggesting it was confounded by technical biases

Which protein was tested as a potential recruiter of proteins into extracellular vesicles (EVs)?

SDCBP (syntenin-1)

What experimental method was used to validate SDCBP's role in EV recruitment?

Knockout of SDCBP in U373vlll cells followed by proteomic analysis of EVs

How many of six SDCBP interaction partners showed significantly reduced EV protein levels upon SDCBP knockout?

Three (CALM1, CEP55, HPRT1)

What is a tissue-preferentially expressed (TiP) gene?

A gene whose expression is enriched in one or a few tissues compared to others

How many tissue-specific PPI networks were derived from HuRI, and how many PPIs does each contain on average?

35 tissue networks, each containing about 25,000 PPIs

What is the dominant interaction pattern between TiP and non-TiP proteins in tissue networks?

Extensive interactions between TiP and non-TiP proteins, but few TiP-TiP PPIs

In how many of the 35 tissues are TiP proteins significantly close to each other in the network?

Only six of 35 tissues

What explains the enrichment of TiP-TiP proximity in four of those six tissues?

Clusters of specifically expressed keratins (KRT) or late-cornified envelope (LCE) proteins

What does the paper conclude about how tissue-specific functions arise?

TiP proteins adapt basic cellular processes by interacting with uniformly expressed proteins, rather than forming isolated TiP-only modules

How does the fraction of tissue-specific PPIs in HuRI compare to the fraction of tissue-specific genes in the genome?

The fraction of tissue-specific PPIs is higher than the fraction of tissue-specific genes

What is a key limitation of Y2H assays mentioned in the paper?

Limited sensitivity; cannot detect PPIs that depend on post-translational processing unique to human cells or require additional partners for stabilization

Where can the HuRI data be accessed?

At http://interactome-atlas.org and through IntAct with identifier IM-25472

What is the name of the final HuRI dataset version published in 2020?

HI-III-20 (Human Interactome obtained from screening Space III, published in 2020)

How many PPIs are in the union of HuRI and all previous CCSB systematic screens (HI-union)?

64,006 binary PPIs involving 9,094 proteins

What orthogonal assays were used to validate Y2H-derived PPIs?

MAPPIT (mammalian protein-protein interaction trap) and GPCA (Gaussia princeps complementation assay)

According to the paper, what role do weaker and more transient PPIs play in the interactome?

They may dominate the protein interactome, which explains why most HuRI PPIs were found in only one scr