Polyklonaler Kaninchen-Antikörper Anti-CRBN

Cereblon (CRBN) was originally identified as a protein involved in human intellectual disability and has recently been recognized as a negative regulator of adenosine monophosphate-activated protein kinase (AMPK)  in vivo  and  in vitro  .  Here we present results showing that CRBN can effectively regulate new protein synthesis via the mammalian target rapamycin (mTOR) signaling pathway, a downstream target of AMPK. While Crbn deficiency suppressed protein translation by activating the AMPK-mTOR cascade in  Crbn -knockout mice, ectopic expression of wild-type CRBN increased protein synthesis by inhibiting endogenous AMPK.

In contrast to wild-type CRBN, a mutant CRBN missing the last 24 amino acids found in human patients failed to abolish mTOR-dependent repression of protein synthesis in Crbn-deficient mouse fibroblasts. These results provide the first evidence that Crbn can activate the protein synthesis machinery via the mTOR signaling pathway by inhibiting AMPK. Given that mTOR-regulated protein synthesis is essential for various forms of synaptic plasticity underlying brain cognitive functions, the results of this study suggest a plausible mechanism for the involvement of CRBN in higher brain functions in humans, and they might also help explain how a specific mutation occurs in  CRBNcan impair the patient’s cognitive abilities.

introduction

Cereblon  (  CRBN  ),  a gene on human chromosome 3p26.2, was originally described as a candidate gene for a mild form of autosomal recessive nonsyndromic mental retardation (ARNSMR). Subsequently, the CRBN protein was characterized in several different cellular contexts. CRBN interacts with the cytoplasmic region of high-conductance calcium-activated potassium channels (  _BKCa  ) to regulate surface expression of the channel protein (Figure). Furthermore, CRBN is the primary target of thalidomide-induced teratogenicity and is thought to besubstrate receptor of an E3 ubiquitin ligase complex (Figure ). A recent study showed that CRBN interacts with the α-subunit of adenosine monophosphate-activated protein kinase (AMPK) and   inhibits  AMPK activation both in vitro  and  in vivo.

• AMPK, a key sensor of cellular energy balance, increases ATP-producing catabolic pathways and inhibits ATP-consuming anabolic pathways. AMPK, a serine/threonine protein kinase, is a heterotrimer composed of a catalytic α subunit and two regulatory subunits, β and γ. AMPK activity can be modulated by phosphorylation at a threonine residue (Thr-172) by upstream kinases such as liver kinase B1 (LKB1).
•  AMPK activation inhibits energy-consuming anabolic processes such as protein translation and achieves these effects largely through inhibition of mammalian target-of-rapamycin (mTOR) signaling.
• The conserved serine-threonine protein kinase mTOR regulates cell growth, proliferation, and synaptic plasticity by controlling protein synthesis. Activation of mTOR acts on one of the primary triggers for the initiation of Cap-dependent translation through the phosphorylation and activation of S6 kinase (S6K1) and through the phosphorylation and inactivation of a repressor of mRNA translation, the eukaryotic initiation factor 4E binding protein ( 4E-BP1).
• Two biochemically distinct mTOR complexes, mTORC1 and mTORC2, are found in mammalian cells, and the activity of mTORC1 is regulated by AMPK. AMPK can repress mTORC1 activity by directly phosphorylating at least two regulatory proteins, tuberous sclerosis 2 (TSC2) and raptor.
• Despite the importance of CBRN for brain function suggested by clinical and experimental evidence, the molecular etiology of the cognitive phenotypes resulting from  CRBN  mutation has not been elucidated. In this study, we investigated the functional role of CRBN as an upstream regulator of mTOR signaling. Our results show that CRBN can upregulate Cap-dependent translation by inhibiting AMPK.
• In contrast to wild-type (WT) CRBN, a mutant CRBN lacking the C-terminal 24 amino acids (R419X) was unable to regulate mTOR signaling due to its inability to repress AMPK activity. Because new protein synthesis is essential for various forms of synaptic plasticity in the brain, defects in the CRBN  -dependent regulation of mTOR signaling may represent the molecular mechanism underlying the learning and memory defects associated with  CRBN  – Underlying mutation.

EXPERIMENTAL PROCEDURES