What is the transmembrane 'Positive-Inside Rule' nowadays? Has the definition changed over time?

Question

First definition.

Two publications by von Heijne in 1989 and 1992 coined the 'Positive-Inside rule' and showed it's practical value in topology prediction of transmembrane helices. It was clearly defined and evidenced that in bacteria the positively charged residues more commonly were found on the "inside" of a membrane (the cytoplasm rather than the periplasm).

Recent Literature.

However it seems the field has moved away from the idea of "inside the compartment" to "inside the cytoplasm" as more data became available. In a 2006 review even von Heijne describes some transmembrane helices as abiding to the positive inside rule because their positively charged residues were inside the cytoplasm, although never explicitly backtracks on the original definition. A similar review in 2007 by von Heijne offers a more definitive refinement of the rule, however removes the concept from being applicable to subcellular membranes.

...the loops connecting the helices differ in amino acid composition, depending on whether they face the inside or outside of the cell (the “positive-inside” rule).

We are now faced with an awkward rule that doesn't account for proteins elsewhere in the secretory pathway or in the other organelles. All those proteins are inside the cell.

More recently still large-scale analysis of transmembrane helices from different biological membrane surfaces, Sharpe et al., 2010 and Baeza-Delgado et al., 2013, show the clustering of positive charge being cytosolic rather than discussing it in terms of the inside of the compartment. Both of these papers still say that they corroborate with the positive inside rule.

Question.

To me it seems that the definition is somewhat sloppy and can broadly be used to say "inside the cytoplasm", despite publications being reluctant to define the rule clearly. Has the definition ever explicitly been changed, or has the field silently changed what "inside" means? ... Or have I completely misinterpreted something? How do the organelles fit into any of these definitions?

Answer 1

I think you have misunderstood the "inside" part of the "positive-inside rule". Perhaps because "inside" is indeed an imprecise term (but now it is history and cannot be changed ;) ). In order to understand it a bit better it helps to think about the topology of the membrane. During synthesis most membrane proteins (ignoring peroxisomal and mitochondrial proteins, which are a whole other topic) are inserted into the ER membrane by the Sec translocon. The parts of the membrane protein that are exposed to the lumen are matured by the ER and Golgi apparatus to be presented on the extracellular surface. Secretory proteins (those that will ultimately be exported from the cell) are made in similar way except they pass entirely through the translocon and end up in the ER lumen. Therefore, the lumen of the ER (and consequently the lumen of other endomembrane compartments) are topologically consistent with the extracellular space, "outside", and not the "inside" of the cell. Perhaps is is easiest to see in a picture (borrowed from here: http://www.bioon.com/book/biology/mboc/chapter12/figure12-4.gif) Coloured in pink are compartments whose topology is consistent with the "outside" and coloured grey are compartments whose topology is consistent with the "inside". So, for membrane proteins found in the endosome, for example, amino acid residues in the lumen are actually "outside" not "inside". It is true that this may not be immediately clear to those not steeped deeply in the field. As the initial studies used bacteria, which do not have membrane-enclosed subcellular compartments, "inside" was simple and made sense. Perhaps "positive-in-the-cytoplasm rule" would have been more accurate. Hope this helps!