Diaz-Meco M T, Municio M M, Sanchez P, Lozano J, Moscat J
Centro de Biología Molecular Severo Ochoa, (Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid), Universidad Autónoma, Spain.
Mol Cell Biol. 1996 Jan;16(1):105-14. doi: 10.1128/MCB.16.1.105.
The members of the atypical subfamily of protein kinase C (PKC) show dramatic structural and functional differences from other PKC isotypes. Thus, in contrast to the classical or novel PKCs, they are not activated by diacylglycerol or phorbol esters. However, the atypical PKCs are the target of important lipid second messengers such as ceramide, phosphatidic acid, and 3'-phosphoinositides. The catalytic and pseudosubstrate sequences in the two atypical PKCs (lambda/iota PKC and zeta PKC) are identical but are significantly different from those of conventional or novel PKCs. It has been shown that microinjection of a peptide with the sequence of the pseudosubstrate of the atypical PKC isotypes but not of alpha PKC or epsilon PKC dramatically inhibited maturation and NF-kappa B activation in Xenopus oocytes, as well as reinitiation of DNA synthesis in quiescent mouse fibroblasts. This indicates that either or both atypical isoforms are important in cell signalling. Besides the pseudosubstrate, the major differences in the sequence between lambda/iota PKC and zeta PKC are located in the regulatory domain. Therefore, any functional divergence between the two types of atypical PKCs will presumably reside in that region. We report here the molecular characterization of lambda-interacting protein (LIP), a novel protein that specifically interacts with the zinc finger of lambda/iota PKC but not zeta PKC. We show in this paper that this interaction is detected not only in vitro but also in vivo, that LIP activates lambda/iota PKC but not zeta PKC in vitro and in vivo, and that this interaction is functionally relevant. Thus, expression of LIP leads to the transactivation of a kappa B-dependent promoter in a manner that is dependent on lambda/iota PKC. To our knowledge, this is the first report on the cloning and characterization of a protein activator of a PKC that binds to the zinc finger domain, which has so far been considered a site for binding of lipid modulators. The fact that LIP binds to lambda/iota PKC but not to the highly related zeta PKC isoform suggests that the specificity of the activation of the members of the different PKC subfamilies will most probably be accounted for by proteins like LIP rather than by lipid activators.
蛋白激酶C（PKC）非典型亚家族的成员与其他PKC同工型在结构和功能上存在显著差异。因此，与经典或新型PKC不同，它们不会被二酰甘油或佛波酯激活。然而，非典型PKC是重要脂质第二信使（如神经酰胺、磷脂酸和3'-磷酸肌醇）的作用靶点。两种非典型PKC（λ/ιPKC和ζPKC）中的催化序列和假底物序列相同，但与传统或新型PKC的序列有显著差异。研究表明，显微注射具有非典型PKC同工型假底物序列的肽（而非αPKC或εPKC的假底物序列）能显著抑制非洲爪蟾卵母细胞的成熟和NF-κB激活，以及静止小鼠成纤维细胞中DNA合成的重新启动。这表明一种或两种非典型同工型在细胞信号传导中都很重要。除了假底物外，λ/ιPKC和ζPKC之间序列的主要差异位于调节结构域。因此，这两种非典型PKC之间的任何功能差异可能都存在于该区域。我们在此报告λ相互作用蛋白（LIP）的分子特征，LIP是一种新型蛋白，它能特异性地与λ/ιPKC的锌指结构域相互作用，而不与ζPKC相互作用。我们在本文中表明，这种相互作用不仅在体外能检测到，在体内也能检测到；LIP在体外和体内都能激活λ/ιPKC而不是ζPKC；并且这种相互作用在功能上是相关的。因此，LIP的表达以依赖于λ/ιPKC的方式导致κB依赖性启动子的反式激活。据我们所知，这是关于一种与锌指结构域结合的PKC蛋白激活剂的克隆和特征的首次报道，迄今为止，锌指结构域一直被认为是脂质调节剂的结合位点。LIP与λ/ιPKC结合但不与高度相关的ζPKC同工型结合这一事实表明，不同PKC亚家族成员激活的特异性很可能是由像LIP这样的蛋白而非脂质激活剂决定的。
