The Ins and Outs of Clusterin: Its Role in Cancer, Eye Diseases and Wound Healing

Figures

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  Figure 3. Generation of sCLU and nCLU. psCLU is transported into the rough ER by the leader signaling peptide (1), and then undergoes cleavage and extensive glycosylation while being transported to the Golgi apparatus (2). The result is a secreted protein of 75–80 kDa with five disulphide bonds between the α- and β-subunits that is secreted outside the cell (3). pnCLU does not undergo any cleavage or extensive glycosylation, and resides in the cytoplasm of unstressed cells. It becomes the mature form (55 kDa) once it is transported to the nucleus (see legend to Figure 2 for the color code) (adapted from ref [55]).

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  Figure 1. CLU gene and transcripts. Schematic representation of CLU gene organization on human chromosome 8 and of the three main mRNA variants. Green blocks represent remaining exons 2–9. Gray blocks represent unique exon 1 of the different CLU transcripts (isoform 1 NM_001831.4: exon 1a with 5′-extended sequence in dark gray, isoform 2 NR_038335.2: 1b and isoform 3 NR_045494.1: 1c). Transcriptional start sites are represented with ATG (or AUG in mRNA). Alternative splicing of variant 1 pre-mRNA generates an mRNA (isoform 1 [Δex2] [23]) that lacks exon 2 and the leader peptide sequence (black box). The position of the sCLU start codon (framed) is defined as nt = 1. Notice the additional in-frame AUG codons on exon 3 of all mRNAs and on exon 1c of variant 3. The canonical transcription start site of CLU mRNA isoform 1 is located at the begining and 23 nucleotides downstream of the TATA promoter element. The termination codon UGA is located in exon 9 at position 12,328 nt.

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  Figure 2. (A) Predicted translation product of the most abundant transcript (isoform 1), and (B) schematic representation of human psCLU protein precursor structure (top) and mature protein (bottom). The precursor polypeptide chain (top) is cleaved proteolytically to remove the 22-mer secretory signal peptide (pink) and subsequently cleaved (black arrow) between Arg 227 and Ser 228 to generate the α (yellow) and β (light blue) chains. These are assembled in anti-parallel orientation to give a heterodimeric molecule (bottom) in which the cysteine-rich centers (brown) are linked by five disulfide bonds (brown ellipses in panels (A,B)) and are flanked by two predicted coiled-coil α-helices (green) and three predicted amphipathic α-helices (dark blue). The six sites of N-linked glycosylation are indicated as yellow stars. nCLU contains 3 nuclear localization signals (NLS) (black) that relocate the nCLU protein from the cytoplasm to the nucleus. The predicted secreted protein is a 449-amino-acids sequence (adapted from ref. [41]).

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  Figure 4. Main regulatory mechanisms of CLU expression. CLU gene can be regulated in different ways. DNA is coiled around histone proteins, allowing it to be efficiently compacted. The modulation of CLU gene expression is closely associated with specific histone modifications. Permissive histone marks, such as H3K9Ac, H3K9S10P and H3K4me3, as well as repressive marks, such as H3K27me3 and H3K9me3, present on the CLU promotor region regulate CLU transcription. The addition of methyl groups to the CpG island that alter transcriptional activity without any changes to the DNA sequence is also another CLU regulatory mechanism. Methylation of the CLU promoter region (black dots) is associated with weak CLU protein expression, while hypomethylation (white dots) allows CLU gene expression. Transcriptional control of the CLU gene transcription can also involve key transcription factors that bind to specific regulatory DNA sequences present on the CLU gene promoter to either repress (repressors; red bars) or induce (activators; green arrows) CLU gene transcription. In addition, microRNAs (miRNAs) are small single-stranded, non-coding RNAs that post-transcriptionally regulate mRNA expression by…

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  Figure 5. CLU isoforms and functions (see legend to Figure 2 for the color code of CLU proteins).

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  Figure 6. Anatomy and structure of the human eye.