The myotubularins are a large family of lipid phosphatases with specificity 
towards PtdIns3P and PtdIns(3,5)P(2). Each of the 14 family members bears a 
signature phosphatase domain, which is inactive in six cases due to amino acid 
changes at the catalytic site. Fragmentary data have indicated heteromeric 
interactions between myotubularins, which have hitherto paired an active family 
member with an inactive one. In this study we have conducted a largescale 
analysis of potential associations within the human myotubularin family, through 
directed two-hybrid screening and immunoprecipitation of epitope-tagged 
proteins. We have confirmed all previously reported combinations and identified 
novel heteromeric interactions: MTMR8 with MTMR9, and MTMR3 with MTMR4, the 
first such combination of enzymatically active MTMs. We also report the capacity 
of several family members to self-associate, including MTMR3 and MTMR4. 
Subcellular localisation studies reveal a unique distribution of MTMR4 to 
endosomal structures, the major site of substrate lipid accumulation. All active 
MTMs we have tested (MTM1, MTMR2-MTMR4) reduce endosomal PtdIns3P levels upon 
overexpression. Despite this, only MTMR4 exerts any effect on EGF receptor 
trafficking and degradation, which is more pronounced with a phosphatase 
inactive form of MTMR4 and requires an intact FYVE domain.