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Objective: Batten disease (BD), the juvenile form of neuronal ceroid lipofuscinosis (NCLs), is pathological characterized by finding lysosomal storage of autofluorescent lipofuscins with unique ultrastructural profiles. The gene underlying BD is designated CLN_3 and encodes a protein, Battenin, of unknown function that localizes in lysosomes and/or mitochondria. Previously, we hypothesized that Battenin associates with other membrane protein(s) to form a membrane complex. Dysfunction of this complex could result in the pathological changes of BD, and possibly in other NCLs. Two such membranous proteins, the slow and fast Battenin-interactive proteins (BIP_s and BIP_f) of unknown functions, have been identified. In this study, we have characterized the functional domains of Battenin that interact with both BIP proteins. Methods: Protein-protein interactions with a yeast two-hybrid system were employed. A “deletion assay” was employed to localize the interactive segment(s). Different lengths of cDNA sequences lacking exon 1-5 were used to express CLN_3-encoded proteins lacking N-terminal segments in the yeast two-hybrid system. N-terminal exons of CLN_3 were deleted with PCR-cloning strategies.Results: We eliminated the possibility of interacting domains from the exon 7-encoded region because both Battenin and mBattenin interact with the BIP proteins. We have shown that peptide sequences encoded by exons 2 and 4 of CLN_3 gene include the functional domains by which Battenin interacts with the BIP proteins. Conclusion: Our studies provide evidence that the N-terminus of Battenin is the functional domain for these protein interactions.
Objective: Batten disease (BD), the juvenile form of neuronal ceroid lipofuscinosis (NCLs), is pathological characterized by finding lysosomal storage of autofluorescent lipofuscins with unique ultrastructural profiles. The gene underlying BD is designated CLN_3 and encodes a protein, Battenin, of unknown function that localizes in lysosomes and / or mitochondria. Previously, we hypothesized that Battenin associates with other membrane protein (s) to form a membrane complex. Dysfunction of this complex could result in the pathological changes of BD, and possibly in other NCLs. Two such membranous proteins, the slow and fast Battenin-interactive proteins (BIP_s and BIP_f) of unknown functions, have been identified. In this study, we have characterized the functional domains of Battenin that interact with both BIP proteins. Methods: Protein-protein interactions with a yeast two-hybrid system were employed. A “deletion assay” was employed to localize the interactive segment (s). Different lengths of cDNA sequences lacking exon 1-5 were used to express CLN_3-encoded proteins lacking N-terminal segments in the yeast two-hybrid system. N-terminal exons of CLN_3 were deleted with PCR-cloning strategies. Results: We eliminated likely of interacting domains from the exon 7-encoded region because both Battenin and mBattenin interact with the BIP proteins. We have shown that the peptide sequences encoded by exons 2 and 4 of the CLN 3 genes include the functional domains by which Batten interacts with the BIP proteins. : Our studies provide evidence that the N-terminus of Battenin is the functional domain for these protein interactions.