Phosphoinositides (PIs) play essential functions as signalling molecules, either directly or by acting as precursors to other signalling molecules. Although their functions have been mostly elucidated in the cytoplasm, PIs are also intra‐nuclear where they contribute to chromatin remodelling and mRNA processing. In addition, PI‐specific kinases, phosphatases as well as phospholipases regulate PI levels within the nucleus. In a previous study we identified ErbB3 binding protein 1 (Ebp1) as a nuclear PI‐binding protein by lipid pull‐down and mass spectrometry. Ebp1 is a conserved protein located in the cytoplasm and nucleus and its differential localisation is correlated to its role in cell proliferation. EBP1 is known to have different cellular activities such as proliferation, differentiation and apoptosis depending on the expression of its long form p48 or the shorter form p42. In this study, we have validated that recombinant p48‐Ebp1 binds to different sets of PIs via two distinct PI‐binding sites located at the N‐ and C‐termini and site‐directed mutagenesis revealed that these PI‐binding sites consist of patches of basic residues. The C‐terminal PI interaction motif was shown to contribute to the sub‐cellular targeting of EBP1 to the nucleolus. We have hence also determined that endogenous EBP1 colocalises with a marker of the nucleolus, nucleophosmin, as well as the class I PI3K catalytic subunit p110β and its product phosphatidylinositol(3,4,5)triphosphate in the nucleolus. Mutations in the same PI‐interaction motif have been identified in different carcinomas and one of these mutations prevents nucleolar targeting. Taken together, these results suggest that PIs may retain Ebp1 in different sub‐cellular compartments to affect its functions.