# Evidence for Rab7b and Its Splice Isoforms Having Distinct Biological Functions from Rab7a

## TL;DR

This study shows that Rab7b and its splice variants have different functions compared to Rab7a, challenging the assumption that they behave similarly.

## Contribution

The paper provides the first direct comparison of Rab7a and Rab7b, revealing distinct functional differences and novel splice isoforms of Rab7b.

## Key findings

- Rab7a and Rab7b nucleotide mutants showed significant differences in their behavior.
- Rab7b splice isoforms caused vesicle aggregation with a distinct phenotype.
- Rab7b vesicles preferentially localized to the Trans Golgi Network, unlike Rab7a.

## Abstract

The Rab family of small guanosine triphosphatases (GTPases) are nucleotide-dependent switches. Mutations in Rabs can result in human diseases. Rab7a and Rab7b transition from early endosomes to lysosomes and are presumed to function similarly. Most studies look at Rab7a, less on Rab7b, with the underlying assumption they function similarly. There have yet to be articles comparing them side by side. Whilst cloning Rab7 homologues, we identified splice isoforms for Rab7b only. These splice isoforms, Rab7b2 and Rab7bx8 lacking different exons, have not been previously characterized but suggest alternative function(s) for Rab7b. Thus, we hypothesize that Rab7 homologues have distinct functions. Here, we compare Rab7a and Rab7b nucleotide mutants locked in GDP-bound (Rab7T22N), GTP-bound (Rab7Q67L), nucleotide-free (Rab7aN125I/Rab7bN124I) states and characterized localization of the Rab7b splice isoforms. HeLa cells were transiently transfected with fluorescently tagged Rab7 reporters. Confocal images were processed with ImageJ and analyzed with SPSS. Rab7a and Rab7b nucleotide mutants were significantly different to one another. Approximately 50% of Rab7b splice isoform-expressing cells had aggregated vesicles, which were phenotypically different from Rab7b vesicles. Rab7a and Rab7b vesicles shared approximately 60% colocalization with each other, while Rab7b vesicles preferentially localized to the Trans Golgi Network. Our results suggest Rab7b is distinct from Rab7a, and Rab7b splice isoforms have different biological functions.