Role of the ubiquitin-like modifier FAT10 in protein degradation and ...

Chapter 2
teins which contain intrinsic targeting signals (Asher et al., 2006). A diverse
group of proteins has been described which can be characterized by the presence
of one or more loosely folded domains and which can be degraded by the 20S pro-
teasome without the need for prior ubiquitylation or ATP-dependent unfolding.
Members of this group include IκBα (Alvarez-Castelao and Castaño, 2005), p53
(Asher et al., 2005), p21 Waf1/Cip1 (Touitou et al., 2001), α-synuclein (Tofaris et al.,
2001), thymidylate synthase (Forsthoefel et al., 2004), and ODC (Asher et al.,
2005). As many of these proteins function as part of a multimeric complex, and
“default” degradation can only be observed when they are present in monomeric
form, it has been suggested that assembly into a larger complex masks the un-
structured regions – which are considered to function as the targeting signal –
and thereby protects them from degradation. It is interesting to note that most,
if not all of these proteins can also be actively targeted to the 26S proteasome by
attachment of a polyubiquitin chain or – in the case of ODC – by interaction with
AZ.
The structure of FAT10 remains yet to be solved, however, the high sequence sim-
ilarity to ubiquitin and ISG15 suggests that it is in all likelihood composed of two
ubiquitin-fold domains (Narasimhan et al., 2005), which are anything but loosely
folded. One can thus safely assume that FAT10 has evolved as a component of
a specialized regulatory system responsible for targeting a subset of proteins for
proteasomal degradation, and is not merely degraded due to an intrinsic instabil-
ity. This notion is further supported by the finding that purified 20S proteasome
– which was fully competent in the cleavage of short peptide substrates (data not
shown) – was unable to degrade HA-FAT10-DHFR in vitro (Fig. 15C), and that
C-terminal attachment of a FAT10-tag does not promote degradation (data not
shown).
The precise mechanism by which FAT10 is targeted to the proteasome has some-
what remained a mystery. Discovery of the UBL-UBA domain protein NUB1L as
a non-covalent interaction partner and an accelerator of FAT10 degradation (Hipp
et al., 2004) suggested that it might interact with FAT10 through its C-terminal
UBA domains and with the 19S regulator through its N-terminal UBL domain
and thus physically link FAT10 to the proteasome. It was soon revealed, however,
that while NUB1L did interact with FAT10 through all three of its UBA domains,
only the UBL domain was required to accelerate its degradation, and that FAT10
was perfectly able to interact with the 26S proteasome on its own (Schmidtke
et al., 2006). Recent studies investigating other members of this family, most
notably Rad23/hHR23 and Dsk2/hPLIC, have revealed functional redundancy in
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