Funktionelle Analysen zu Erkrankungen, die zu einer Dysfunktion/Fehlregulation ribosomaler Proteine führen

Functional analyses to diseases which lead to dysfuction/dysregulation of ribosomal proteins

by Miriam D. Noceti
Doctoral thesis
Date of Examination:2023-01-19
Date of issue:2023-01-20
Advisor:Prof. Dr. Silke J. Pauli
Referee:Prof. Dr. Silke J. Pauli
Referee:Prof. Dr. Christof Kramm
Persistent Address: http://dx.doi.org/10.53846/goediss-9666

 

 

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Abstract

English

Ribosomopathies are characterized by impaired synthesis or function of ribosomes. Mutations in genes which code for components of the normal ribosomal biogenesis lead to these rare congenital diseases. Functional analyses of the clinical pictures of two patients should reveal if the underlying genetic mutations are associated with a dysfunction or dysregulation of ribosomal proteins. Diamond Blackfan Anemia (DBA) with symptoms of erythrocyte aplasia and association to cancer is one of the more common ribosomopathies. Pathogenic variants in more than 19 ribosomal protein (RP) genes are known as causative. In a patient with the clinical diagnosis of DBA and increased incidence of tumor diseases in his family, a variant of uncertain significance (VUS) in the ribosomal protein RPS27A has been detected. RPS27A is known to interact with MDM2 in the RP-MDM2-p53-pathway. In this study, the VUS in RPS27A revealed no significant impact on the expression of RPS27A on RNA or protein levels in different cell types. Also, the variant showed no influence on the RP-MDM2-p53-pathway. An indirect interaction between RPS27A and MDM2 could be confirmed. This suggests that other interaction proteins must interact with RPS27A and MDM2 on which the RPS27A variant could have an impact. The second patient had symptoms similar to two children with variants in FBRSL1 but had instead a microdeletion in 22q12.2-q12.2 that includes 32 genes, one of them is ZMAT5. FBRSL1 is a paralogue of AUTS2, which is regulated by ZMAT3, and both belong to the same Polycomb group complex. In the case of a downregulation of FBRSL1, different ribosomal proteins are also downregulated. In this study it was shown that ZMAT5 is expressed on RNA and protein level in different cells. Moreover, qRT-PCRs revealed a downregulation of ZMAT5 in patient fibroblasts. Performing an siRNA knockdown of ZMAT5 a significant downregulation of the long isoform of FBRSL1 containing an AUTS2-domain in patient fibroblasts could be demonstrated, while no impact of the heterozygous ZMAT5 deletion on the expression of different RP genes could be detected. Because a dysregulation of RP genes in blood cells has been found, tissue specific differences are discussed. In conclusion, a regulatory loop between ZMAT5 and one isoform of FBRSL1 was found, but classification of the two clinical pictures as ribosomopathies has still to be proven.
Keywords: ZMAT5; Ribosomopathies; FBRSL1; RPS27A; Diamond Blackfan Anemia; Ribosomal proteins; Polycomb complex protein