The inherited blindness protein AIPL1 regulates the ubiquitin-like FAT10 pathway

Mutations in AIPL1 cause the inherited blindness Leber congenital amaurosis (LCA). AIPL1 has previously been shown to interact with NUB1, which facilitates the proteasomal degradation of proteins modified with the ubiquitin-like protein FAT10. Here we report that AIPL1 binds non-covalently to free FAT10 and FAT10ylated proteins and can form a ternary complex with FAT10 and NUB1. In addition, AIPL1 antagonised the NUB1-mediated degradation of the model FAT10 conjugate, FAT10-DHFR, and pathogenic mutations of AIPL1 were defective in inhibiting this degradation. While all AIPL1 mutants tested still bound FAT10-DHFR, there was a close correlation between the ability of the mutants to interact with NUB1 and their ability to prevent NUB1-mediated degradation. Interestingly, AIPL1 also co-immunoprecipitated the E1 activating enzyme for FAT10, UBA6, suggesting AIPL1 may have a role in directly regulating the FAT10 conjugation machinery. These studies are the first to implicate FAT10 in retinal cell biology and LCA pathogenesis, and reveal a new role of AIPL1 in regulating the FAT10 pathway.     

The Leber congenital amaurosis protein AIPL1 modulates the nuclear translocation of NUB1 and suppresses inclusion formation by NUB1 fragments

Mutations in the aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) cause the blinding disease Leber congenital amaurosis (LCA). The similarity of AIPL1 to AIP has led to suggestions that AIPL1 could function in a similar manner to AIP in facilitating protein translocation and as a component of chaperone complexes. AIPL1 interacts with the cell cycle regulator NEDD8 ultimate buster protein 1 (NUB1). As AIPL1 is predominantly cytoplasmic and NUB1 is predominantly nuclear, we tested the hypothesis that AIPL1 could modulate the nuclear translocation of NUB1. Co-transfection of AIPL1 with GFP-NUB1 resulted in a shift of GFP-NUB1 subcellular distribution toward the cytoplasm. Interestingly, AIPL1 was able to act in a chaperone-like fashion to efficiently suppress inclusion formation by NUB1 fragments. Co-transfection of AIPL1 with GFP-NUB1-N and GFP-NUB1-C resulted in an AIPL1-dependent suppression of GFP-NUB1-N perinuclear inclusions and GFP-NUB1-C intranuclear inclusions leading to the redistribution of these fragments in the cytoplasm. This chaperone-like function of AIPL1 was specific for NUB1, since AIPL1 was unable to suppress the inclusion formation by unrelated aggregation-prone proteins and AIP had no effect on NUB1 localization or inclusion formation. We examined the effect of a range of pathogenic and engineered mutations on the ability of AIPL1 to modulate NUB1 localization or inclusion formation. With the exception of W278X, which formed non-functional SDS-insoluble inclusions, all of the pathogenic mutations studied were soluble and could modulate NUB1 with varying efficiency compared with the wild-type protein. The effect of AIPL1 on NUB1 required the C-terminal region of AIPL1, as engineered C-terminal truncation mutations had no effect on NUB1. These data show that AIPL1 can modulate protein translocation and act in a chaperone-like manner and suggest that AIPL1 is an important modulator of NUB1 cellular function.     

NEDD8 ultimate buster-1L interacts with the ubiquitin-like protein FAT10 and accelerates its degradation

FAT10 is an interferon-gamma-inducible ubiquitin-like protein that consists of two ubiquitin-like domains. FAT10 bears a diglycine motif at its C terminus that can form isopeptide bonds to so far unidentified target proteins. Recently we found that FAT10 and its conjugates are rapidly degraded by the proteasome and that the N-terminal fusion of FAT10 to a long lived protein markedly reduces its half-life. FAT10 may hence direct target proteins to the proteasome for degradation. In this study we report a new interaction partner of FAT10 that may link FAT10 to the proteasome. A yeast two-hybrid screen identified NEDD8 ultimate buster-1L (NUB1L) as a non-covalent binding partner of FAT10, and this interaction was confirmed by coimmunoprecipitation and glutathione S-transferase pull-down experiments. NUB1L is also an interferon-inducible protein that has been reported to interact with the ubiquitin-like protein NEDD8, thus leading to accelerated NEDD8 degradation. Here we show that NUB1L binds to FAT10 much stronger than to NEDD8 and that NEDD8 cannot compete with FAT10 for NUB1L binding. The interaction of FAT10 and NUB1L is specific as green fluorescent fusion proteins containing ubiquitin or SUMO-1 do not bind to NUB1L. The coexpression of NUB1L enhanced the degradation rate of FAT10 8-fold, whereas NEDD8 degradation was only accelerated 2-fold. Because NUB1 was shown to bind to the proteasome subunit RPN10 in vitro and to be contained in 26 S proteasome preparations, it may function as a linker that targets FAT10 for degradation by the proteasome.     

NUB1-mediated targeting of the ubiquitin precursor UbC1 for its C-terminal hydrolysis.

NEDD8 is a ubiquitin-like protein that controls vital biological events through its conjugation to target proteins. Previously, we identified a negative regulator of the NEDD8 conjugation system, NEDD8 ultimate buster-1 (NUB1), that recruits NEDD8 and its conjugates to the proteasome for degradation. Recently, we performed yeast two-hybrid screening with NUB1 as bait and isolated a ubiquitin precursor UbC1 that is composed of nine tandem repeats of a ubiquitin unit through alpha-peptide bonds. Interestingly, NUB1 interacted with UbC1 through its UBA domain. Further study revealed that the UBA domain interacted with alpha-peptide bond-linked polyubiquitin, but not with isopeptide bond-linked polyubiquitin, indicating that the UBA domain of NUB1 is a specific acceptor for the linear ubiquitin precursor. A functional study revealed that an unidentified protein that was immunoprecipitated with NUB1 served as a ubiquitin C-terminal hydrolase for UbC1. Thus, NUB1 seems to form a protein complex with the unidentified ubiquitin C-terminal hydrolase and recruit UbC1 to this complex. This might allow the ubiquitin C-terminal hydrolase to hydrolyze UbC1, in order to generate ubiquitin monomers. Northern blot analysis showed that the mRNAs of both NUB1 and UbC1 were enriched in the testis. Furthermore, in situ hybridization showed that both mRNAs were strongly expressed in seminiferous tubules of the testis. These results may imply that the UbC1 hydrolysis mediated by NUB1 is involved in cellular functions in the seminiferous tubules such as spermatogenesis.     

Interaction of NUB1 with the proteasome subunit S5a

NUB1 interacts with a ubiquitin-like protein NEDD8 to target the NEDD8 monomer and neddylated proteins to the proteasome for degradation. Therefore, NUB1 is thought to be a potent downregulator of NEDD8 conjugation system. Since NUB1 possesses a UBL domain, which was previously shown to be an S5a-interacting motif in RAD23/HHR23, we initially hypothesized that NUB1 interacts with the S5a subunit of the proteasome through its UBL domain. To examine this, we performed an in vitro GST pull-down assay and a yeast two-hybrid assay. Unexpectedly, our studies revealed that NUB1 directly interacts with the S5a subunit through its C-terminal region between amino acid residues 536 and 584, not through its UBL domain. Although the UBL domain was not an S5a-interacting motif in NUB1, our further studies revealed that the UBL domain is required for the function of NUB1.     

Degradation of FAT10 by the 26S proteasome is independent of ubiquitylation but relies on NUB1L

The ubiquitin-like modifier FAT10 targets proteins for degradation by the proteasome, a process accelerated by the UBL-UBA domain protein NEDD8 ultimate buster 1-long. Here, we show that FAT10-mediated degradation occurs independently of poly-ubiquitylation as purified 26S proteasome readily degraded FAT10-dihydrofolate reductase (DHFR) but not ubiquitin-DHFR in vitro. Interestingly, the 26S proteasome could only degrade FAT10-DHFR when NUB1L was present. Knock-down of NUB1L attenuated the degradation of FAT10-DHFR in intact cells suggesting that NUB1L determines the degradation rate of FAT10-linked proteins. In conclusion, our data establish FAT10 as a ubiquitin-independent but NUB1L-dependent targeting signal for proteasomal degradation.     

Nuclear-cytoplasmic interaction in chlorophyll-deficient soybean,Glycine max (Fabaceae)

In higher plants, plastids and mitochondria are the predominant carriers of extrachromosomal genetic information. There is interplay between the plastids, the mitochondria, and the nuclear genome. In soybean, Glycine max (L.) Merr., both nuclearly and maternally inherited chlorophyll-deficient mutants have been described. Conditional lethality previously was reported in soybean when maternally inherited chlorophyll-deficient mutant (Genetic Type T275) was crossed with nuclearly inherited yellow foliar malate dehydrogenase null mutants (Genetic Types T253 and T323). Our objective was to test for conditional lethality when maternally inherited yellow foliar mutants T278, T314, T315, T316, T319, and T320 were female parents and nuclearly inherited yellow foliar malate dehydrogenase null mutants T253 and T323 were male parents. Our results indicated conditional lethality in the F₂ generation when any of the six cytoplasmically inherited yellow foliar mutants were female parents and either T253 or T323 were male parents. The physiological nature of conditional lethality is not known. Data indicate a common basis in soybean for conditional lethality among the cytoplasmically inherited yellow foliar mutants when crossed with the nuclearly inherited yellow foliar malate dehydrogenase null mutants. No interactions were observed between cytoplasmically inherited or nuclearly inherited green seed embryo mutants as female parents and either T253 or T323 as male parents.     

Photoproduction of hydrogen by sulfur-deprived <Emphasis Type="Italic">C. reinhardtii</Emphasis> mutants with impaired Photosystem II photochemical activity

Photoproduction of H₂ was examined in a series of sulfur-deprived Chlamydomonas reinhardtii D1-R323 mutants with progressively impaired PSII photochemical activity. In the R323H, R323D, and R323E D1 mutants, replacement of arginine affects photosystem II (PSII) function, as demonstrated by progressive decreases in O₂-evolving activity and loss of PSII photochemical activity. Significant changes in PSII activity were found when the arginine residue was replaced by negatively charged amino acid residues (R323D and R323E). However, the R323H (positively charged or neutral, depending on the ambient pH) mutant had minimal changes in PSII activity. The R323H, R323D, and R323E mutants and the pseudo-wild-type (pWt) with restored PSII function were used to study the effects of sulfur deprivation on H₂-production activity. All of these mutants exhibited significant changes in the normal parameters associated with the H₂-photoproduction process, such as a shorter aerobic phase, lower accumulation of starch, a prolonged anaerobic phase observed before the onset of H₂-production, a shorter duration of H₂-production, lower H₂ yields compared to the pWt control, and slightly higher production of dark fermentation products such as acetate and formate. The more compromised the PSII photochemical activity, the more dramatic was the effect of sulfur deprivation on the H₂-production process, which depends both on the presence of residual PSII activity and the amount of stored starch.     

Effect of capsular polysaccharide of Klebsiella pneumoniae on host resistance to bacterial infections. I. Induction of increased susceptibility to infections in mice.

When Klebsiella pneumoniae capsular polysaccharide (CPS-K) from type 1, Kasuya strain, was injected intraperitoneally (i.p.) immediately before i.p. bacterial challenge, the survival time of mice infected with Salmonella enteritidis NUB 1 (virulent strain) was shortened and the mortality rate for mice infected with S. enteritidis NUB 31 (avirulent strain) was enhanced. The promotion of infection with S. enteritidis NUB 1 by CPS-K depended upon its dose, the effect of CPS-K being demonstrable up to as little as 0.2 mug per mouse. In the case of S. enteritidis NUB 31, the effect of CPS-K was detectable only when more than 20 mug per mouse was injected. As a result of enumeration of bacterial populations in the peritoneal washing, blood, liver and spleen, it was revealed that CPS-K promoted in vivo growth of S. enteritidis NUB 1 and NUB 31. In addition, CPS-K enhanced the mortality rate in mice infected with Streptococcus pyogenes or Streptococcus pneumoniae. The peak CPS-K effect on infection with S. enteritidis NUB 1 was seen when given immediately before bacterial challenge. The active substance responsible for the infection-promoting effect of CPS-K was neutral CPS-K, which is distinct from the O antigen and from acidic CPS-K (the type-specific capsular antigen). Preparations of neutral CPS-K isolated from the other three strains of K. pneumoniae exhibited a marked infection-promoting effect comparable with that of preparations from the Kasuya strain. Neutral CPS-K, with identical antigenicity to that from the Kasuya strain, has already been found to exert a strong adjuvant effect on antibody responses to various antigens in mice. No parallelism exists between infection-promoting activity and adjuvant activity of neutral CPS-K.     

Regulation of the NEDD8 conjugation system by a splicing variant,NUB1L

NEDD8 is a ubiquitin-like protein that controls vital biological events through its conjugation to target proteins. We previously identified a negative regulator of the NEDD8 conjugation system, NUB1, which works by recruiting NEDD8 and its conjugates to the proteasome for degradation. Recently, we found its splicing variant, NUB1L. It possesses an insertion of 14 amino acids that codes for a UBA domain. Structural study revealed that NUB1 has a NEDD8-binding site at the C terminus, whereas NUB1L has an additional site at the newly generated UBA domain. Interestingly, the sequence A(X4)L(X10)L(X3)L was conserved in these NEDD8-binding sites among human and other mammals. Mutational studies revealed that at least three Leu residues in the conserved sequence are required for binding with NEDD8. Functional study suggested that the NEDD8-binding ability at the C terminus of NUB1 and NUB1L is mainly involved in the down-regulation of NEDD8, but the NEDD8-binding ability at the UBA2 domain of NUB1L is minimally or not involved at all. The NEDD8-binding ability at the UBA2 domain might be required for an unknown function of NUB1L.     

Effect of Capsular Polysaccharide of Klebsiella pneumoniae on Host Resistance to Bacterial Infections

When Klebsiella pneumoniae capsular polysaccharide (CPS-K) from type 1, Kasuya strain, was injected intraperitoneally (i.p.) immediately before i.p. bacterial challenge, the survival time of mice infected with Salmonella enteritidis NUB 1 (virulent strain) was shortened and the mortality rate for mice infected with S. enteritidis NUB 31 (avirulent strain) was enhanced. The promotion of infection with S. enteritidis NUB 1 by CPS-K depended upon its dose, the effect of CPS-K being demonstrable up to as little as 0.2 μg per mouse. In the case of S. enteritidis NUB 31, the effect of CPS-K was detectable only when more than 20 μg per mouse was injected. As a result of enumeration of bacterial populations in the peritoneal washing, blood, liver and spleen, it was revealed that CPS-K promoted in vivo growth of S. enteritidis NUB 1 and NUB 31. In addition, CPS-K enhanced the mortality rate in mice infected with Streptococcus pyogenes or Streptococcus pneumoniae. The peak CPS-K effect on infection with S. enteritidis NUB 1 was seen when given immediately before bacterial challenge. The active substance responsible for the infection-promoting effect of CPS-K was neutral CPS-K, which is distinct from the O antigen and from acidic CPS-K (the type-specific capsular antigen). Preparations of neutral CPS-K isolated from the other three strains of K. pneumoniae exhibited a marked infection-promoting effect comparable with that of preparations from the Kasuya strain. Neutral CPS-K, with identical antigenicity to that from the Kasuya strain, has already been found to exert a strong adjuvant effect on antibody responses to various antigens in mice. No parallelism exists between infection-promoting activity and adjuvant activity of neutral CPS-K.     

NEDD8 Ultimate Buster 1 Long (NUB1L) Protein Suppresses Atypical Neddylation and Promotes the Proteasomal Degradation of Misfolded Proteins

Neddylation is a posttranslational modification that controls diverse biological processes by covalently conjugating the ubiquitin-like protein NEDD8 to specific targets. Neddylation is commonly mediated by NEDD8-specific enzymes (typical neddylation) and, sometimes, by ubiquitin enzymes (atypical neddylation). Although typical neddylation is known to regulate protein function in many ways, the regulatory mechanisms and biological consequence of atypical neddylation remain largely unexplored. Here we report that NEDD8 conjugates were accumulated in the diseased hearts from mouse models and human patients. Proteotoxic stresses induced typical and atypical neddylation in cardiomyocytes. Loss of NUB1L exaggerated atypical neddylation, whereas NUB1L overexpression repressed atypical neddylation through promoting the degradation of NEDD8. Activation of atypical neddylation accumulated a surrogate misfolded protein, GFPu. In contrast, suppression of atypical neddylation by NUB1L overexpression enhanced GFPu degradation. Moreover, NUB1L depletion accumulated a cardiomyopathy-linked misfolded protein, CryAB^R120G, whereas NUB1L overexpression promoted its degradation through suppressing neddylation of ubiquitinated proteins in cardiomyocytes. Consequently, NUB1L protected cells from proteotoxic stress-induced cell injury. In summary, these data indicate that NUB1L suppresses atypical neddylation and promotes the degradation of misfolded proteins by the proteasome. Our findings also suggest that induction of NUB1L could potentially become a novel therapeutic strategy for diseases with increased proteotoxic stress.     

Mutation or deletion of the C-terminal tail affects the function and structure of Xenopus laevis small heat shock protein,hsp30

Small heat shock proteins (shsps) act as molecular chaperones by preventing heat-induced aggregation and unfolding of cellular proteins by a mechanism that is still unclear. Previously we found that the C-terminal end of Xenopus shsp, hsp30C (30C), was essential for optimal chaperone activity. Examination of the C-terminal tail of 30C revealed that it had a net negative charge. Involvement of this negative charge in chaperone activity was assessed by the creation of two mutants, D209G (Asp converted to the more neutrally charged and less polar Gly at position 209) and D209/213G (Asp to Gly at position 209 and 213). Compared to 30C and D209G, D209/213G was impaired in inhibiting heat-induced citrate synthase aggregation. In rabbit reticulocyte lysate and Xenopus oocyte microinjection refolding assays the mutants were not as efficient as 30C in maintaining heat-treated luciferase in a folding competent state. Circular dichroism analysis revealed that D209G was similar in secondary structure to 30C whereas D209/213G displayed a loss of alpha-helical-like and beta-sheet structure. Also, C-terminal truncation of 30C or 30D (an hsp30 isoform) resulted in a loss of secondary structure and function. This study clearly shows that mutation of aspartic acid residues in the C-terminal end of hsp30 or its truncation disrupts secondary structure and impairs its chaperone activity.     

In Rhodobacter sphaeroides reaction centers, mutation of proline L209 to aromatic residues in the vicinity of a water channel alters the dynamic coupling between electron and proton transfer processes.

The X-ray crystallographic structure of the photosynthetic reaction center from Rhodobacter sphaeroides obtained at high resolution has revealed a number of internal water molecules (Ermler, U., Fritzsch, G., Buchanan, S. K., and Michel, H. (1994) Structure 2, 925-936; Stowell, M. H. B., McPhillips, T. M., Rees, D. C., Soltis, S. M., Abresch, E., and Feher, G. (1997) Science 276, 812-816). Some of them are organized into distinct hydrogen-bonded water chains that connect Q(B) (the terminal quinone electron acceptor of the reaction center) to the aqueous phase. To investigate the role of the water chains in the proton conduction process, proline L209, located immediately adjacent to a water chain, was mutated to the following residues: F, Y, W, E, and T. We have first analyzed the effects of the mutations on the kinetic and thermodynamic properties of the rate constants of the second electron transfer (k(AB)(2)) and of the coupled proton uptake (k(H)+) at the second flash. In all aromatic mutants, k(AB)(2) and k(H)+ are notably and concomitantly decreased compared to the wild-type, while no effect is observed in the other mutants. The temperature dependence of these rates shows activation energy values (DeltaH) similar for the proton and electron-transfer processes in the wild-type and in most of the mutants, except for the L209PW and L209PF mutants. The analysis of the enthalpy factors related to the electron and proton-transfer processes in the L209PF and the L209PW mutants allows to distinguish the respective effects of the mutations for both transfer reactions. It is noteworthy that in the aromatic mutants a substantial increase of the free energies of activation is observed (DeltaG(L209PY) < DeltaG(L209PF) < DeltaG(L209PW)) for both proton and electron-transfer reactions, while in the other mutants, DeltaG is not affected. The salt concentration dependence of k(AB)(2) shows, in the L209PF and L209PW mutants, a higher screening of the protein surface potential experienced by Q(B). Our data suggest that residues F and W in position L209 increase the polarizability of the internal water molecules and polar residues by altering the organization of the hydrogen-bond network. We have also analyzed the rates of the first electron-transfer reaction (k(AB)(1)), in the 100 micros time domain. These kinetics have previously been shown to reflect protein relaxation events possibly including proton uptake events (Tiede, D. M., Vazquez, J., Cordova, J., and Marone, P. M. (1996) Biochemistry 35, 10763-10775). Interestingly, in the L209PF and L209PW mutants, k(AB)(1) is notably decreased in comparison to the wild type and the other mutants, in a similar way as k(AB)(2) and k(H)+. Our data imply that the dynamic organization of this web is tightly coupled to the electron transfer process that is kinetically limited by protonation events and/or conformational rearrangements within the protein.     

Cellular characteristics of neuroblastoma cells: regulation by the ELR--CXC chemokine CXCL10 and expression of a CXCR3-like receptor

Bone marrow stroma cells secrete the chemokine CXCL12 that may support bone marrow metastasis formation by neuroblastoma cells. The present study demonstrates that bone marrow stroma cell lines also secrete CXCL10, a chemokine that was shown in the past to have anti-malignancy functions. A receptor recognized by antibodies against CXCR3 was shown to be expressed by six neuroblastoma cell lines. Further detailed analysis was performed on the NUB6 and SK-NMC neuroblastoma cells, showing that CXCL10 induced potent Erk phosphorylation in a G(alpha)i-dependent manner. The role of a CXCR3-like receptor in Erk phosphorylation was substantiated by the ability of CXCL11, another potent CXCR3 ligand, to induce Erk phosphorylation in the NUB6 and SK-NMC cells. Further characterization of CXCL10 activities indicated that CXCL10 partly inhibited the growth of the NUB6 and SK-NMC cells. Both NUB6 and SK-NMC cells did not migrate to CXCL10, although their migratory machinery was intact, as evidenced by their migration to bone marrow constituents. Altogether, these results suggest that CXCL10 interacts with a CXCR3-like receptor in neuroblastoma cell lines, raising the possibility that following the homing of the tumor cells to the bone marrow (through a CXCL10-independent mechanism), CXCL10 may partly inhibit neuroblastoma cell growth at this site.     

NEDD8 Ultimate Buster-1 Long (NUB1L) Protein Promotes Transfer of NEDD8 to Proteasome for Degradation through the P97UFD1/NPL4 Complex

The NEDD8 protein and neddylation levels in cells are modulated by NUB1L or NUB1 through proteasomal degradation, but the underlying molecular mechanism is not well understood. Here, we report that NUB1L down-regulated the protein levels of NEDD8 and neddylation through specifically recognizing NEDD8 and P97/VCP. NUB1L directly interacted with NEDD8, but not with ubiquitin, on the key residue Asn-51 of NEDD8 and with P97/VCP on its positively charged VCP binding motif. In coordination with the P97-UFD1-NPL4 complex (P97^UFD1/NPL4), NUB1L promotes transfer of NEDD8 to proteasome for degradation. This mechanism is also exemplified by the canonical neddylation of cullin 1 for SCF (SKP1-cullin1-F-box) ubiquitin E3 ligases that is exquisitely regulated by the turnover of NEDD8.     

The UBA domains of NUB1L are required for binding but not for accelerated degradation of the ubiquitin-like modifier FAT10

Proteins selected for degradation are labeled with multiple molecules of ubiquitin and are subsequently cleaved by the 26 S proteasome. A family of proteins containing at least one ubiquitin-associated (UBA) domain and one ubiquitin-like (UBL) domain have been shown to act as soluble ubiquitin receptors of the 26 S proteasome and introduce a new level of specificity into the degradation system. They bind ubiquitylated proteins via their UBA domains and the 26 S proteasome via their UBL domain and facilitate the contact between substrate and protease. NEDD8 ultimate buster-1 long (NUB1L) belongs to this class of proteins and contains one UBL and three UBA domains. We recently reported that NUB1L interacts with the ubiquitin-like modifier FAT10 and accelerates its degradation and that of its conjugates. Here we show that a deletion mutant of NUB1L lacking the UBL domain is still able to bind FAT10 but not the proteasome and no longer accelerates FAT10 degradation. A version of NUB1L lacking all three UBA domains, on the other hand, looses the ability to bind FAT10 but is still able to interact with the proteasome and accelerates the degradation of FAT10. The degradation of a FAT10 mutant containing only the C-terminal UBL domain is also still accelerated by NUB1L, even though the two proteins do not interact. In addition, we show that FAT10 and either one of its UBL domains alone can interact directly with the 26 S proteasome. We propose that NUB1L not only acts as a linker between the 26 S proteasome and ubiquitin-like proteins, but also as a facilitator of proteasomal degradation.